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 examined the expression of the cyclin-dependent kinase 4, p34PSK-J3/cdk4 protein, in small dense, activated, and proliferating primary B lymphocytes. A small steady state level of cdk4 synthesis was detected in resting B cells. Stimulation of resting B cells with mitogenic amounts of F(ab')2 fragments of goat anti-mouse IgM (anti-Ig) resulted in increased synthesis of cdk4 protein during the mid to late G1 phase of the cell cycle; LPS or the combination of phorbol ester and calcium ionophore also elevated cdk4 levels. Resting B cells that we rendered competent by treatment with IL 4 or low doses of anti-Ig or, alternatively, were activated by phorbol ester or ionomycin alone also exhibited heightened cdk4 protein levels. Subsequent analysis of potential cdk4 regulatory subunit D-type cyclins revealed that cyclin D2, not cyclin D1 or D3, is expressed in primary mature B lymphocytes. The induction of cyclin D2 synthesis in response to mitogenic anti-Ig paralleled cdk4 expression; however, IL-4 or low dose anti-Ig alone did not increase the rate of de novo cyclin D2 synthesis above that of resting B cells. The significance of the lack of cyclin D2 regulation by competence-inducing growth factors was demonstrated, in that only mitogenic factors that stimulated DNA synthesis 1) led to the formation of stable cyclin D2/cdk4 holoenzyme complexes during G1 phase progression, and 2) afforded the isolation of anti-cyclin D2 or anti-cdk4 immunoprecipitates that phosphorylated retinoblastoma. These findings suggest a role for these proteins during the mid to late G1 phase progression and possibly the G1/S phase transition in primary mature B lymphocytes.
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PMID:Regulation of the catalytic subunit (p34PSK-J3/cdk4) for the major D-type cyclin in mature B lymphocytes. 854 4

Prostaglandin A2 (PGA2) potently inhibits cell proliferation and suppresses tumor growth in vivo, but little is known regarding the molecular mechanisms mediating these effects. Here we demonstrate that treatment of breast carcinoma MCF-7 cells with PGA2 leads to G1 arrest associated with a dramatic decrease in the levels of cyclin D1 and cyclin-dependent kinase 4 (cdk4) and accompanied by an increase in the expression of p21. We further show that these effects occur independent of cellular p53 status. The decline in cyclin D and cdk4 protein levels is correlated with loss in cdk4 kinase activity, cdk2 activity is also significantly inhibited in PGA2-treated cells, an effect closely associated with the upregulation of p21. Immunoprecipitation experiments verified that p21 was indeed complexed with cdk2 in PGA2-treated cells. Additional experiments with synchronized MCF-7 cultures stimulated with serum revealed that treatment with PGA2 prevents the progression of cells from G1 to S. Accordingly, the kinase activity associated with cdk4, cyclin E, and cdk2 immunocomplexes, which normally increases following serum addition, was unchanged in PGA2-treated cells. Furthermore, the retinoblastoma protein (Rb), a substrate of cdk4 and cdk2 whose phosphorylation is necessary for cell cycle progression, remains underphosphorylated in PGA2-treated serum-stimulated cells. These findings indicate that PGA2 exerts its growth-inhibitory effects through modulation of the expression and/or activity of several key G1 regulatory proteins. Our results highlight the chemotherapeutic potential of PGA2, particularly for suppressing growth of tumors lacking p53 function.
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PMID:Inhibition of G1 cyclin-dependent kinase activity during growth arrest of human breast carcinoma cells by prostaglandin A2. 862 77

In contrast to mature B cells, immature stage B cells do not proliferate following Ag receptor cross-linking with anti-Ig Abs. To determine where in the cell cycle immature B cells arrest, we have examined the expression of specific G, cell cycle regulators. Following surface IgM (sIgM) cross-linking on mature B cells, we observed increased expression of the early G1 kinase, cyclin-dependent kinase 4 (cdk4), and one of its regulatory subunits, cyclin D2. Mature B cells also showed increased expression of components required for G1/S transition, including cyclin E and cdk2. Whereas immature stage B cells increased expression of cyclin D2 and cdk4 after anti-IgM stimulation, unlike mature stage B cells they failed to express cyclin E and cdk2. Expression of cyclin D2 and cdk4 indicates that these cells can exit G0 and enter the initial G1 phase following sIgM ligation. Interestingly, IL-4, which by itself does not stimulate proliferation of immature B cells, induced expression of cyclin E and cdk2. These latter results suggest that IL-4 complements sIgM, signaling for proliferation by increasing the basal levels of late G1 cell cycle regulators. Consistent with this idea, IL-4 synergizes with anti-Ig Abs to promote cell cycle progression and proliferation of immature B cells. Finally, c-myc, a transcriptional regulator of some members of the cell cycle machinery, is not induced following sIgM cross-linking of immature cells. This lack of inducible expression contrasts with that seen in mature stage B cells, and in immature stage cells stimulated to proliferate with LPS. These results suggest that c-myc may be a component of the signaling pathway that induces cyclin E and cdk2 expression.
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PMID:Immature stage B cells enter but do not progress beyond the early G1 phase of the cell cycle in response to antigen receptor signaling. 864 97

The retinoblastoma protein (Rb) is essential for the maintenance of the postmitotic state in terminally differentiated myocytes. Upon C2C12 myogenesis, the level of the cyclin-dependent kinase 4 (CDK4) protein does not change, but its Rb kinase activity is down-regulated markedly. Here, we show that the reduction in CDK4 activity results from (a) the irreversible induction and association of the p21 CDK inhibitor with the CDK4 complex and (b) a decline in overall D-type cyclin expression. Immunoprecipitation-coupled immunoblot analyses demonstrated that myocyte differentiation produces alterations in the subunit interactions within the CDK4 complex, including a diminished interaction with cyclin D1 and enhanced interactions with cyclin D3 and p21. The significance of the p21 interaction with CDK4 was indicated by the ability of anti-p21 antibodies to specifically immunodeplete a Rb kinase inhibitory activity that was bound to the CDK4 complex in myotubes. Furthermore, the restimulation of myotubes with serum did not lead to the re-activation of CDK4 or disrupt the CDK4-p21 interaction. Despite the increase in cyclin D3 expression during myogenesis, quantitative immunoblot analyses revealed that the combined levels of cyclin D1 and D3 decline during this process and that CDK4 is expressed at much higher levels than either of these cyclin subunits in postmitotic myotubes. These results suggest that the myogenesis-induced up-regulation of p21 and down-regulation of the total D-type cyclin expression contribute to the inhibition of the CDK4 Rb kinase activity, leading to conditions that favor the accumulation of the hypophosphorylated Rb and growth arrest upon terminal differentiation.
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PMID:Inhibition of retinoblastoma protein phosphorylation by myogenesis-induced changes in the subunit composition of the cyclin-dependent kinase 4 complex. 893 Mar 96

The human T-cell leukemia virus type 1 (HTLV-1) Tax oncoprotein causes cellular transformation by deregulating important cellular processes such as DNA repair, transcription, signal transduction, proliferation, and growth. Although it is clear that normal cell cycle control is deregulated during HTLV-1-induced cellular transformation, the effects of Tax on cell cycle control are not well understood. Flow cytometric analyses of human T cells indicate that cell cycle arrest in late G1, at or before the G1/S restriction point, by p16INK4a is relieved by Tax. Furthermore, Tax-dependent stimulation of 5-bromo-2'-deoxyuridine incorporation and transcriptional activation is inhibited by p16INK4a. This result suggests that p16INK4a is able to block Tax-dependent stimulation of DNA synthesis and cell cycle progression into S phase. In vitro binding assays with recombinant glutathione S-transferase fusion proteins and [35S]methionine-labeled proteins indicate that Tax binds specifically with p16INK4a but not with either p21cip1 or p27kip1. Furthermore, sequential immunoprecipitation assays with specific antisera and [35S]methionine-labeled cell lysates subsequent to coexpression with Tax and p16INK4a indicate that the two proteins form complexes in vivo. Immunocomplex kinase assays with cyclin-dependent kinase 4 antiserum indicate that Tax blocks the inhibition of cdk4 kinase activity by p16INK4a. This study identifies p16INK4a as a novel cellular target for Tax and suggests that the inactivation of p16INK4a function is a mechanism of cell cycle deregulation by Tax.
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PMID:Human T-cell leukemia virus type 1 Tax releases cell cycle arrest induced by p16INK4a. 903 27

Basic fibroblast growth factor (bFGF), a classical mitogen in fibroblasts and endothelial cells, inhibits the proliferation of MCF-7 and other human breast cancer cell lines. To explain this paradoxic effect, we investigated the effects of bFGF on cyclins and protein members of cyclin complexes that exert positive and negative control on the progression of cells through the G1 phase of the cell cycle. bFGF induced an increase in cyclin D1, cyclin E, and cyclin-dependent kinase 4 (cdk4) protein levels in a bFGF dose-dependent manner. However, bFGF also induced a heat-stable, transferable cytoplasmic factor in MCF-7 cells that inhibited the histone H1 kinase activity of reconstituted cyclin E-cdk2 and cyclin A-cdk2 complexes from Mv1Lu mink lung epithelial cells. The appearance of this inhibitor correlated with a bFGF dose- and time-dependent increase in the levels of cdk inhibitor p21(WAF1/CIP1) mRNA and protein. The increase in the level of p21(WAF1/CIP1) was associated with the disappearance of the rapidly migrating, activated form of cdk2 from cell lysates, dephosphorylation of the retinoblastoma protein (Rb), and a decrease in cyclin A levels. These changes were represented in the cyclin D1 and E complexes by an increased association with p21(WAF1/CIP1), proliferating cell nuclear antigen (PCNA), and the inactive form of cdk2, without an absolute change in cellular PCNA levels and by a switch in the association of cyclin D1 complexes with the hyperphosphorylated form to the dephosphorylated form of Rb. These experiments demonstrate that stimulation of MCF-7 cells with bFGF, although resulting in up-regulation of G1 proteins responsible for mitogenic events, also induces a concomitant decrease in cyclin A levels and an increase in p21(WAF1/CIP1) mRNA and protein and results in inactivation of cdk2, dephosphorylation of Rb, and a segregation of PCNA to the G1 cyclin complexes. The dual, conflicting signaling by bFGF results in a net inhibitory phenotype in these cells. These experiments suggest a pleiotropic role for bFGF in breast cancer.
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PMID:Basic fibroblast growth factor causes growth arrest in MCF-7 human breast cancer cells while inducing both mitogenic and inhibitory G1 events. 913 19

The involvement of cell cycle-regulatory proteins in apoptosis of neuronally differentiated PC12 cells induced by the removal of nerve growth factor and serum was examined. Three major findings are presented. (1) Cdc2 kinase protein levels increased fivefold in apoptotic PC12 cells by day 3 of serum and nerve growth factor deprivation. Histone H1 kinase activity was increased significantly in p13(suc1) precipitates of apoptotic PC12 cells, which was due to increased activation and/or expression of cdc2 kinase. (2) The protein levels of cyclin-dependent kinase 4, cyclin D, and proliferating cell nuclear antigen that are normally expressed in the cell cycle were increased during neuronal PC12 cell apoptosis. (3) The levels of the catalytic subunit, but not the regulatory subunit of the calcium/calmodulin-dependent protein phosphatase 2B, decreased significantly concomitant with a significant decrease in protein phosphatase 2B activity early in the apoptotic process. Protein phosphatase 2A activity decreased slightly but significantly after 3 days of serum and nerve growth factor deprivation, and no alterations in protein phosphatase 1 were observed during the apoptotic process. These data demonstrate that certain cell cycle-regulatory proteins are inappropriately expressed and that alterations in specific phosphorylation events, as indicated by the increase in histone H1 kinase activity and the decrease in protein phosphatase 2B activity, are most likely occurring during apoptosis of PC12 cells. These observations support the hypothesis that apoptosis may be due in part to a nondividing cell's uncoordinated attempt to reenter and progress through the cell cycle.
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PMID:Select alterations in protein kinases and phosphatases during apoptosis of differentiated PC12 cells. 916 26

Members of the INK4 protein family specifically inhibit cyclin-dependent kinase 4 (cdk4) and cdk6-mediated phosphorylation of the retinoblastoma susceptibility gene product (Rb). p16INK4A, a prototypic INK4 protein, has been identified as a tumor suppressor in many human cancers. Inactivation of p16INK4A in tumors expressing wild-type Rb is thought to be required in order for many malignant cell types to enter S phase efficiently or to escape senescence. Here, we demonstrate another mechanism of tumor suppression by implicating p16INK4A in a G1 arrest checkpoint in response to DNA damage. Calu-1 non-small cell lung cancer cells, which retain Rb and lack p53, do not arrest in G1 following DNA damage. However, engineered expression of p16INK4A at levels compatible with cell proliferation restores a G1 arrest checkpoint in response to treatment with gamma-irradiation, topoisomerase I and II inhibitors, and cisplatin. A similar checkpoint can be demonstrated in p53-/- fibroblasts that express p16INK4A. DNA damage-induced G1 arrest, which requires the expression of pocket proteins such as Rb, can be abrogated by overexpression of cdk4, kinase-inactive cdk4 variants capable of sequestering p16INK4A, or a cdk4 variant incapable of binding p16INK4A. After exposure to DNA-damaging agents, there was no change either in overall levels of p16INK4A or in amounts of p16INK4A found in complex with cdks 4 and 6. Nonetheless, p16INK4A expression is required for the reduction in cdk4- and cdk6-mediated Rb kinase activity observed in response to DNA damage. During tumor progression, loss of p16INK4A expression may be necessary for cells with wild-type Rb to bypass this G1 arrest checkpoint and attain a fully transformed phenotype.
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PMID:p16INK4A participates in a G1 arrest checkpoint in response to DNA damage. 941 85

Mammalian fibroblasts require mitogens in order to exit from G0 (quiescence) and progress through the G1 phase of the cell cycle, although once they pass the restriction point late in G1 they can enter S phase and complete the cell cycle without mitogens [1]. Mitogenic signals are integrated through the GTPase Ras, which regulates the levels of cyclin D1 [2-5], a component of the cell cycle machinery that operates during G1 phase by activating cyclin-dependent kinase 4 (Cdk4). The accumulation of active cyclin E-Cdk2 complexes also requires Ras [6]. These two G1 cyclin-Cdk complexes act on a family of E2F-associated transcriptional repressors typified by the retinoblastoma protein (Rb) to bring about a transcriptional program that promotes passage through S phase [7-9], but can also activate DNA replication independently of Rb-E2F [10-12]. Although G1 cyclin-Cdk complexes are required for S-phase entry and can shorten G1 phase when overexpressed [13-15], it is not known whether they are sufficient for this transition. Here, we report that serum-starved (G0) diploid human fibroblasts initiate DNA synthesis upon microinjection of active G1 cyclin-Cdk complexes, but not upon microinjection of an S-phase cyclin-Cdk complex. These data indicate that G1 Cdk activation is rate-limiting for S-phase entry, and that Cdk activation is likely to be the primary function of growth factor signalling pathways that lead to DNA synthesis.
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PMID:G1 cyclin-dependent kinases are sufficient to initiate DNA synthesis in quiescent human fibroblasts. 942 30

In this report, we studied the effect of phosmidosine, a proline-containing nucleotide on the serum-induced cell cycle progression in human lung fibroblast WI-38 cells. Phosmidosine suppressed S-phase entry and arrested cell cycle progression at the G1 phase. In serum-stimulated cells, phosmidosine did not affect the activation of the mitogen-activated protein kinase cascade. However, phosmidosine inhibited hyperphosphorylation of retinoblastoma (RB) protein by RB-kinases such as cyclin-dependent kinase 4 and cyclin-dependent kinase 2, probably as a result of the inhibition of cyclin D1 expression. Furthermore, in tsFT210 cells, a temperature-sensitive cdc2 mutant isolated from the mouse mammary carcinoma cell line FM3A, phosmidosine, irreversibly inhibited the cell cycle progression at G1 without affecting the G2 to M transition. Phosmidosine acts at an earlier point in G1 compared with mimosine or aphidicolin, well-known cell cycle blockers at the G1-S boundary. Taken together, phosmidosine arrested cells at a specific point between the start point and restriction point in G1 and is a useful drug that may contribute to the understanding of the regulatory mechanisms of G1 progression.
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PMID:Inhibition of cyclin D1 expression and phosphorylation of retinoblastoma protein by phosmidosine, a nucleotide antibiotic. 948 24


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