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

We have previously reported that certain tyrphostins which block EGF-R phosphorylation in cell-free systems fail to do so in intact cells. Nevertheless, we found that this family of tyrphostins inhibits both EGF- and calf serum-induced cell growth and DNA synthesis [Osherov, N.A., Gazit, C., Gilon, and Levitzki, A. (1993). Selective inhibition of the EGF and HER2/Neu receptors by Tyrphostins. J. Biol. Chem. 268, 11134-11142.] Now we show that these tyrphostins exert their inhibitory activity even when added at a time when the cells have already passed their restriction point and receptor activation is no longer necessary. AG555 and AG556 arrest 85% of the cells at late G1, whereas AG490 and AG494 cause cells to arrest at late G1 and during S phase. No arrest occurs during G2 or M phase. Further analysis revealed that these tyrphostins act by inhibiting the activation of the enzyme Cdk2 without affecting its levels or its intrinsic kinase activity. Furthermore, they do not alter the association of Cdk2 to cyclin E or cyclin A or to the inhibitory proteins p21 and p27. These compounds also have no effect on the activating phosphorylation of Cdk2 by Cdk2 activating kinase (CAK) and no effect on the catalytic domain of cdc25 phosphatase. These compounds lead to the accumulation of phosphorylated Cdk2 on tyrosine 15 which is most probably the cause for its inhibition leading to cell cycle arrest at G1/S. A structure-activity relationship study defines a very precise pharmacophore, suggesting a unique molecular target not yet identified and which is most probably involved in the regulation of the tyrosine-phosphorylated state of Cdk2. These compounds represent a new class of cell proliferation blockers whose target is Cdk2 activation.
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PMID:Inhibition of Cdk2 activation by selected tyrphostins causes cell cycle arrest at late G1 and S phase. 963 76

The induction of premature mitosis by okadaic acid (OA) in HeLa cells in S-phase or in G2-phase has been studied using light microscopy, immunofluorescence, and immunochemical techniques. The observations indicate an involvement of a cdc2-independent pathway in these cells. It has been claimed that inhibition of an OA-sensitive phosphatase, possibly of PP1, induces activation of a kinase which is sensitive to staurosporine and Zn2+. This kinase brings about mitosis-specific cytoskeletal rearrangements, chromosome condensation, and nuclear envelope breakdown, inducing a mitosis-like state. However, other mitotic events do not follow. The possibility that this kinase may be a NIMA-like Nek2 kinase is discussed.
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PMID:Cdc2-independent induction of premature mitosis by okadaic acid in HeLa cells. 966 96

Cdc25 phosphatases play key roles in cell cycle progression by activating cyclin-dependent kinases. In human cells, cdc25 proteins are encoded by a multigene family, consisting of cdc25A, cdc25B and cdc25C. While cdc25A plays a crucial role at the G1/S phase transition, cdc25C is involved in the dephosphorylation and activation of the mitotic kinase, cdc2/cyclinB. In addition, cdc25C itself is regulated by cdc2/cyclinB which then creates a positive feedback loop that controls entry into mitosis. In this study we show that the activity of cdc25B appears during late S phase and peaks during G2 phase. Both in vitro and in vivo cdc25B is activated through phosphorylation during S-phase. Using a cell duplication, microinjection assay we show that ablation of cdc25B function by specific antibodies blocks cell cycle progression in Hs68 cells by inhibition of entry into mitosis. Cdc25B function neither plays a role in later stages of mitosis nor for the inititation of DNA replication. These results indicate that cdc25B is a mitotic regulator that might act as a 'starter phosphatase' to initiate the positive feedback loop at the entry into M phase.
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PMID:The cdc25B phosphatase is essential for the G2/M phase transition in human cells. 968 38

The autonomous cell divisions during the early development of Xenopus laevis believed to comprise a universal cell cycle engine. Recent experimental data indicates that the Cdk2-cyclin E kinase is required for the rapid divisions during Xenopus embryogenesis and that the complex is crucial for the transition into mitosis. In the present paper, the activity of Cdk2-cyclin E is incorporated into an existing comprehensive model of the cell cycle engine as an activity operating in parallel with the mitosis promotion factor (MPF) on the phosphatase Cdc25. This introduces interesting regulatory and dynamic properties for the transition into mitosis that reveals new insight into the mechanisms of the cell division process. It is shown that the Cdk2-cyclin E complex can act as an effective modulator of the threshold MPF activity needed to initiate mitosis. When the Cdk2-cyclin E activity is below a critical value, the cell cycle arrests in a well-defined state of low MPF activity corresponding to G2 arrest. In agreement with experiments a single mitotic event occurs following injection of free cyclin B. Above a critical activity, the presence of Cdk2-cyclin E allows for sustained oscillations corresponding to repeated cell divisions and the Cdk2-cyclin E may be the cause for the suppressed G2 checkpoint in the early embryonic cell cycles. A detailed bifurcation analysis reveals that the transition from steady to oscillatory behavior involves a homoclinic orbit of infinite period through an omega explosion. The general properties of the omega explosion explain the bifurcation as a dynamic mechanism well-suited for the G2 checkpoint and suggest a plausible explanation for the elongation of the cell cycle as observed at the mid-blastula transition. The proposed mechanism also suggests a plausible explanation of G2 checkpoint failure following DNA damage in human cells overexpressing Cdk2 and we suggest that the onset of mitosis in the mammalian cell occurs as the result of a slow passage through a critical point.
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PMID:Dynamics of the cell cycle engine: Cdk2-kinase and the transition into mitosis. 968 42

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) inhibits 17beta-estradiol (E2) mammary tumor growth in rodents and in MCF-7 human breast cancer cells; however, the cell cycle genes/proteins which are inhibited have not been determined. Initial studies showed that treatment of MCF-7 cells with 10 nM E2 significantly increased cyclin D1 (protein and mRNA), cdk2- and cdk4-dependent kinase activities, and hyperphosphorylation of retinoblastoma (RB) protein. In contrast to results of recent studies (M. D. Planas-Silva and R. A. Weinberg, 1997, Mol. Cell. Biol. 17, 4059-4069), E2 induced dissociation of both cdk2 and cdk4 proteins from the p21 protein complex and significantly increased cdk7-dependent kinase activity. Treatment of MCF-7 cells with E2 also induced cdc25A phosphatase protein, which was accompanied by increased cdk2 and cdk4 proteins containing unphosphorylated tyrosine residues. Although TCDD alone has minimal effects on cell cycle proteins/enzymes, several E2-induced responses were significantly inhibited in MCF-7 cells cotreated with E2 plus TCDD. For example, TCDD significantly inhibited E2-induced hyperphosphorylation of RB, cyclin D1 protein, and cdk2-, cdk4-, and cdk7-dependent kinase activities. Inhibition of E2-induced cdk4-dependent kinase activity by TCDD may be related to the parallel decrease of E2-induced cyclin D1 protein, and inhibition of induced cdk2- and cdk4-dependent kinase activities may be due to significantly increased p21 levels in cells cotreated with TCDD plus E2. These results demonstrate that the antiestrogenic activity of TCDD is due to downregulation of several E2-induced cell cycle proteins/activities and this illustrates the complex cross talk between the aryl hydrocarbon and the E2 receptor signaling pathways.
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PMID:Aryl hydrocarbon receptor-mediated antiestrogenicity in MCF-7 cells: modulation of hormone-induced cell cycle enzymes. 970 14

Mammalian Cdc25 phosphatase is responsible for the dephosphorylation of Cdc2 and other cyclin-dependent kinases at Thr14 and Tyr15, thus activating the kinase and allowing cell cycle progression. The catalytic domain of this dual-specificity phosphatase has recently been mapped to the 180 most C-terminal amino acids. Apart from a CX3R motif, which is present at the active site of all known tyrosine phosphatases, Cdc25 does not share any obvious sequence similarity with any of those enzymes. Until very recently, the Cdc25 family was the only subfamily of tyrosine phosphates for which no three-dimensional structural data were available. Using the generalized profile technique, a sensitive method for sequence database searches, we found an extended and highly significant sequence similarity between the Cdc25 catalytic domain and similarly sized regions in other proteins: the non-catalytic domain of two distinct families of MAP-kinase phosphates, the non-catalytic domain of several ubiquitin protein hydrolases, the N and C-terminal domain of rhodanese, and a large and heterogeneous groups of stress-response proteins from all phyla. The relationship of Cdc25 to the structurally well-characterized rhodanese spans the entire catalytic domain and served as template for a structural model for human Cdc25a, which is fundamentally different from previously suggested models for Cdc25 catalytic domain organization. The surface positioning of subfamily-specific conserved residues allows us to predict the sites of interaction with Cdk2, a physiological target of Cdc25a. Based on the results of this analysis, we also predict that the budding yeast arsenate resistance protein Acr2 and the ORF Ygr203w encode protein phosphatases with catalytic properties similar to that of the Cdc25 family. Recent determination of the crystal structure of the Cdc25a catalytic domain supports the validity of the model and demonstrates the power of the generalized sequence profile technique in homology-based modeling of the three-dimensional structure of a protein having a weak but significant sequence similarity with a structurally characterized protein.
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PMID:A model of Cdc25 phosphatase catalytic domain and Cdk-interaction surface based on the presence of a rhodanese homology domain. 973 50

Ste20p from Saccharomyces cerevisiae is a member of the Ste20/p21-activated protein kinase family of protein kinases. The Ste20p kinase is post-translationally modified by phosphorylation in a cell cycle-dependent manner, as judged by the appearance of phosphatase-sensitive species with reduced mobility on SDS-polyacrylamide gel electrophoresis. This modification is maximal during S phase, and correlates with the accumulation of Ste20p fused to green fluorescent protein at the site of bud emergence. Overexpression of Cln2p, but not Clb2p or Clb5p, causes a quantitative shift of Ste20p to the reduced mobility form, and this shift is dependent on Cdc28p activity. The post-translational mobility shift can be generated in vitro by incubation of Ste20p with immunoprecipitated Cln2p kinase complexes, but not by immunoprecipitated Clb2p or Clb5p kinase complexes. Ste20p is therefore a substrate for the Cdc28p kinase, and undergoes a Cln2p-Cdc28p mediated mobility shift as cells initiate budding and DNA replication. In cells that express only the Cln2p G1 cyclin, minor overexpression of Ste20p causes aberrant morphology, suggesting a proper coordination of Ste20p and Cln-Cdc28p activity may be required for the control of cell shape.
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PMID:Cell cycle- and Cln2p-Cdc28p-dependent phosphorylation of the yeast Ste20p protein kinase. 977 29

Organization of intermediate filament, a major component of cytoskeleton, is regulated by protein phosphorylation/dephosphorylation, which is a dynamic process governed by a balance between the activities of involved protein kinases and phosphatases. Blocking dephosphorylation by protein phosphatase inhibitors such as okadaic acid (OA) leads to an apparent activation of protein kinase(s) and to genuine activation of phosphatase-regulated protein kinase(s). Treatment of 9L rat brain tumor cells with OA results in a drastically increased phosphorylation of vimentin, an intermediate filament protein. In-gel renaturing assays and in vitro kinase assays using vimentin as the exogenous substrate indicate that certain protein kinase(s) is activated in OA-treated cells. With specific protein kinase inhibitors, we show the possible involvement of the cdc2 kinase- and p38 mitogen-activated protein kinase (p38MAPK)-mediated pathways in this process. Subsequent in vitro assays demonstrate that vimentin may serve as an excellent substrate for MAPK-activated protein kinase-2 (MAPKAPK-2), the downstream effector of p38MAPK, and that MAPKAPK-2 is activated with OA treatment. Comparative analysis of tryptic phosphopeptide maps also indicates that corresponding phosphopeptides emerged in vimentin from OA-treated cells and were phosphorylated by MAPKAPK-2. Taken together, the results clearly demonstrate that MAPKAPK-2 may function as a vimentin kinase in vitro and in vivo. These findings shed new light on the possible involvement of the p38MAPK signaling cascade, via MAPKAPK-2, in the maintenance of integrity and possible physiological regulation of intermediate filaments.
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PMID:Identification of mitogen-activated protein kinase-activated protein kinase-2 as a vimentin kinase activated by okadaic acid in 9L rat brain tumor cells. 977 16

When exposed to diverse growth conditions in vitro, cells can respond by entering states of proliferation, quiescence, differentiation or apoptosis. While the choices among these states can be influenced by proto-oncogene expression, how these disparate outcomes are achieved remains poorly understood. To address these issues, we have generated rodent fibroblast cell lines that harbor a human c-myc gene under the control of a tetracycline-regulated promoter. When Myc-induced cells are deprived of serum growth factors, they rapidly become apoptotic with the onset of apoptosis preceded by a large, transient increase in cdk2 kinase activity that is associated with the induction of cdc25A phosphatase and the later accumulation of p27Kip1 kinase inhibitor. Surprisingly, serum starvation in the absence of myc overexpression, (which leads to quiescence instead of apoptosis) also causes a marked transient elevation in cdk2 kinase activity, an induction of cdc25A and a delayed increase in p27Kip1. Transient elevations in cdk2 kinase activity and cdc25A abundance are required for cell cycle progression, but it is evident that these changes also precede entry to either apoptosis or quiescence in serum-starved cells. These findings suggest that the pathways to both quiescence and apoptosis share regulatory machinery with cell cycle control mechanisms. In addition, the abundance of Myc protein can be critical in the choices among these cellular states.
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PMID:Quiescence versus apoptosis: Myc abundance determines pathway of exit from the cell cycle. 979 26

Exposure of CV-1P cells to hypoxic conditions results in reversible cell cycle arrest concomitant with accumulation of pRB in the hypophosphorylated, growth suppressive form. Similar to cell cycle arrest induced by serum starvation, we show here that hypoxia-induced arrest is accompanied by a decrease in pRB-directed CDK4 and CDK2 activities, lower cyclin D and E protein levels, and by an increase in p27 protein abundance. Immunoprecipitation studies reveal an increase in p27 association with cyclin E-CDK2 complexes. In contrast to cell cycle arrest induced by serum starvation, hypoxia increases PP1-mediated pRB dephosphorylation. These data reveal that synergy between decreased pRB-directed cyclin/CDK activity and increased pRB-directed phosphatase activity contribute towards inducing and maintaining pRB in its hypophosphorylated, growth suppressive state during hypoxia.
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PMID:Hypoxia-induced pRB hypophosphorylation results from downregulation of CDK and upregulation of PP1 activities. 981 60


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