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)

DNA polymerase alpha-primase is known to be phosphorylated in human and yeast cells in a cell cycle-dependent manner on the p180 and p68 subunits. Here we show that phosphorylation of purified human DNA polymerase alpha-primase by purified cyclin A/cdk2 in vitro reduced its ability to initiate simian virus 40 (SV40) DNA replication in vitro, while phosphorylation by cyclin E/cdk2 stimulated its initiation activity. Tryptic phosphopeptide mapping revealed a family of p68 peptides that was modified well by cyclin A/cdk2 and poorly by cyclin E/cdk2. The p180 phosphopeptides were identical with both kinases. By mass spectrometry, the p68 peptide family was identified as residues 141 to 160. Cyclin A/cdk2- and cyclin A/cdc2-modified p68 also displayed a phosphorylation-dependent shift to slower electrophoretic mobility. Mutation of the four putative phosphorylation sites within p68 peptide residues 141 to 160 prevented its phosphorylation by cyclin A/cdk2 and the inhibition of replication activity. Phosphopeptide maps of the p68 subunit of DNA polymerase alpha-primase from human cells, synchronized and labeled in G1/S and in G2, revealed a cyclin E/cdk2-like pattern in G1/S and a cyclin A/cdk2-like pattern in G2. The slower-electrophoretic-mobility form of p68 was absent in human cells in G1/S and appeared as the cells entered G2/M. Consistent with this, the ability of DNA polymerase alpha-primase isolated from synchronized human cells to initiate SV40 replication was maximal in G1/S, decreased as the cells completed S phase, and reached a minimum in G2/M. These results suggest that the replication activity of DNA polymerase alpha-primase in human cells is regulated by phosphorylation in a cell cycle-dependent manner.
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PMID:Cell cycle-dependent regulation of human DNA polymerase alpha-primase activity by phosphorylation. 985 88

Cyclin-dependent protein kinases (Cdks) are key regulatory proteins of the eukaryotic cell cycle. Cdc2 is expressed in late G1/S phase and functions in the G2 to M phase transition. Adenovirus E1A proteins are known to induce the expression of p34cdc2 and DNA synthesis in normal quiescent cells. In this study, mutational analysis of the human cdc2 promoter revealed that transactivation of the promoter by the E1A proteins in cycling cells is mediated through the two CCAAT box binding motifs. A 110-kDa protein (CBF/cdc2) was identified in nuclear extracts from monkey kidney (CV-1) cells stably expressing E1A as well as from adenovirus-transformed human 293 cells. Further, we show that this EIA-inducible CBF/cdc2 is related to the CBF which was shown to activate the heat shock protein 70 promoter. Analyses of the functional domain(s) of E1A required for the induction of the CBF and transactivation of the cdc2 promoter in these conditions revealed that E1A mutants which were defective in binding the pRB family of proteins or the cellular p300 protein were still active in assays measuring the induction of the CBF and transactivation of the cdc2 promoter, albeit with reduced efficiencies. But the E1A mutant which lost both functional domains was inactive in these assays. These results suggest that E1A has redundant functional domains for the induction of the 110-kDa CBF and activation of human cdc2 gene expression.
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PMID:Transactivation of the human cdc2 promoter by adenovirus E1A in cycling cells is mediated by induction of a 110-kDa CCAAT-box-binding factor. 987 26

Deepwater rice (Oryza sativa) is adapted to survive conditions of severe flooding over extended periods of time. During such periods adventitious roots develop to provide water, nutrients, and anchorage. In the present study the growth of adventitious roots was induced by treatment with ethylene but not auxin, cytokinin, or gibberellin. Root elongation was enhanced between 8 and 10 h after submergence. The population of cells in the S phase and expression of the S-phase-specific histone H3 gene increased within 4 to 6 h. Within 6 to 8 h the G2-phase population increased. Cell-cycle activation was accompanied by sequential induction of a cdc2-activating kinase homolog, R2, of two cdc2 genes, cdc2Os-1 and cdc2Os-2, and of three cyclin genes, cycA1;3, cycB2;1, and cycB2;2, but only induction of the R2 gene expression preceded the induction of the S phase, possibly contributing to cell-cycle regulation in the G1 phase. Both cdc2 genes were expressed at slightly higher levels during DNA replication. Transcripts of the A-type cyclin accumulated during the S and G2 phases, and transcripts of the B-type cyclins accumulated during the G2 phase. Cyclin expression was induced at all nodes with a similar time course, suggesting that ethylene acts systemically and that root primordia respond to ethylene at an early developmental stage.
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PMID:Adventitious root growth and cell-cycle induction in deepwater rice 988 Mar 42

Partial hepatectomy (PH) triggers the entry of rat liver cells into the cell cycle. The signals leading to cell-cycle activation converge into a family of kinases named cyclin-dependent kinases (cdks). Specific cyclin-cdk complexes are sequentially activated during the cell cycle. Cyclin D-cdk4 and cyclin E-cdk2 are activated during the G1 phase, cyclin A-cdk2 is activated during the S phase, and cyclin B-cdk1 during mitosis. In the present study, we have examined the timing of the activation of cdk4 and cdk2, the intracellular location of G1/S cyclins and cdks, and the relationship between location and cdk4 and cdk2 activities during rat liver regeneration after a PH. Results showed that the activity of both kinases started at 13 hours and showed maximal levels at 24 hours after hepatectomy. In quiescent cells, cyclin D3 and cdk4 were cytoplasmatic, whereas cyclin D1 was nuclear. At 5 hours after hepatectomy, cyclin D3 and cdk4 began to move into the nucleus, and at 13 hours, they were mostly nuclear. During the first 13 hours after hepatectomy, significant amounts of cyclin D1-cdk4 and cyclin D3-cdk4 complexes were formed, but they were mostly inactive. At 24 hours, these complexes were maximally activated. This activation was associated with the accumulation of cyclin D1, cyclin D3, and cdk4 in a nuclear subfraction extractable with nucleases. At 28 hours, the activity of cdk4 in this nuclear subfraction decreased when cyclin D1 moved from this fraction to the nuclear matrix (NM) and the levels of cyclin D3 diminished. The maximal activation of cdk2 at 24 hours was also associated with the accumulation of cyclin E, cyclin A, and cdk2 in this nuclease-sensitive fraction. The inactivation of cdk2 at 28 hours was associated with a strong decrease in cdk2 in this nuclear subfraction. Thus, results reported here indicate that the activation of cdk4 and cdk2 observed in rat liver cells after a PH is associated with a specific intranuclear location of these cdks and their associated cyclins.
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PMID:Activation of cdk4 and cdk2 during rat liver regeneration is associated with intranuclear rearrangements of cyclin-cdk complexes. 991 14

Proliferation and apoptosis are increased in many types of inflammatory diseases. A role for the cyclin kinase inhibitor p27(Kip1) (p27) in limiting proliferation has been shown. In this study, we show that p27(-/-) mesangial cells and fibroblasts have strikingly elevated rates of apoptosis, not proliferation, when deprived of growth factors. Apoptosis was rescued by restoration of p27 expression. Cyclin A-cyclin-dependent kinase 2 (CDK2) activity, but not cyclin E-CDK2 activity, was increased in serum-starved p27(-/-) cells, and decreasing CDK2 activity, either pharmacologically (Roscovitine) or by a dominant-negative mutant, inhibited apoptosis. Our results show that a new biological function for the CDK inhibitor p27 is protection of cells from apoptosis by constraining CDK2 activity. These results suggest that CDK inhibitors are necessary for coordinating the cell cycle and cell-death programs so that cell viability is maintained during exit from the cell cycle.
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PMID:Modulation of apoptosis by the cyclin-dependent kinase inhibitor p27(Kip1). 1007 76

B-myb is a highly conserved member of the myb proto-oncogene family that encodes a ubiquitously expressed 110-kDa sequence-specific DNA-binding protein. Transactivation of Myb-inducible promoters by B-Myb is repressed by a regulatory domain located at the C-terminus of the protein. Cyclin A/Cdk2-mediated phosphorylation apparently releases the negative constraint and triggers B-Myb transactivation potential. Two-dimensional tryptic phosphopeptide analysis indicated that the majority of the sites phosphorylated in vivo are targeted in vitro by cyclin A/Cdk2. Six sites in B-Myb fulfil the requirements for recognition by Cdk2. Using point mutation of the phosphorylation sites to nonphosphorylatable amino acids, we show that five of these sites are targets for Cdk2 in vivo. Mutation of one of these residues (T524) to alanine diminished the ability of B-Myb to promote transcription of a reporter gene, suggesting that phosphorylation of B-Myb at this site is important for the regulation of its activity by cyclin A/Cdk2.
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PMID:Identification of cyclin A/Cdk2 phosphorylation sites in B-Myb. 1009 72

The G2-M transition of the cell cycle is under the control of the M-phase promoting factor (MPF) formed of cdc2 kinase and cyclin B. The Xenopus prophase-blocked oocyte contains a stockpile of cyclin B2-cdc2 complexes that are maintained inactive by a double inhibitory phosphorylation on Thr-14 and Tyr-15 of cdc2. Free cdc2 molecules that are not associated with cyclin, are present in excess as compared to cyclin B2-associated cdc2. This pool of free cdc2 is permanently recruited to associate with neosynthetized cyclin B2 in the resting prophase oocyte, to feed up the pre-MPF stockpile. During re-entry into meiosis, free cdc2 could generate with newly synthesized cyclin B a small level of active MPF, that could serve as starter to initiate the conversion of pre-MPF into MPF. It was, therefore, of high interest to investigate whether free cdc2 interacts with other proteins and what could be its intracellular localization. To address these questions, we developed an in vitro system of membrane vesicles. We demonstrate here that free cdc2 is recovered in association with the external layer of membrane vesicles, whereas cyclin B2-associated cdc2 is not. Cyclin is able to associate in vitro with cdc2-containing membrane vesicles. This association does not induce the inhibitory cdc2 phosphorylations. However, it does not lead to active complexes, suggesting that membrane vesicles prevent cdc2 activation. C-Raf1, another kinase activated during reentry into meiosis, is also totally recovered in association with the membrane vesicles.
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PMID:In vitro binding of free cdc2 and raf kinase to membrane vesicles: a possible new regulatory mechanism for cdc2 kinase activation in Xenopus oocyte. 1020 51

Cyclin-dependent kinases (Cdks) control major transitions as cells pass through the cell cycle. It has recently been shown that centrosome duplication in vertebrates requires Cdk2 activity and can be driven solely by Cdk2-cyclin E complexes.
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PMID:Cell cycle: driving the centrosome cycle. 1037 18

Both estradiol binding and phosphorylation regulate transcriptional activation by the human estrogen receptor alpha (ER). We have previously shown that activation of the cyclin A-CDK2 complex by overexpression of cyclin A leads to enhanced ER-dependent transcriptional activation and that the cyclin A-CDK2 complex phosphorylates the ER N-terminal activation function-1 (AF-1) between residues 82 and 121. Within ER AF-1, serines 104, 106, and 118 represent potential CDK phosphorylation sites, and in this current study, we ascertain their importance in mediating cyclin A-CDK2-dependent enhancement of ER transcriptional activity. Cyclin A overexpression does not enhance transcriptional activation by an ER derivative bearing serine-to-alanine changes at residues 104, 106, and 118. Likewise, the cyclin A-CDK2 complex does not phosphorylate this triple-mutated derivative in vitro. Individual serine-to-alanine mutations at residues 104 and 106, but not 118, decrease ER-dependent transcriptional enhancement in response to cyclin A. The same relationship holds for ER phosphorylation by cyclin A-CDK2 in vitro. Finally, enhancement of ER transcriptional activation by cyclin A is evident in the absence and presence of estradiol, as well as in the presence of tamoxifen, suggesting that the effect of the cyclin A-CDK2 on ER transcriptional activation is AF-2-independent. These results indicate that the enhancement of ER transcriptional activation by the cyclin A-CDK2 complex is mediated via the AF-1 domain by phosphorylation of serines 104 and 106. We propose that these residues control ER AF-1 activity in response to signals that affect cyclin A-CDK2 function.
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PMID:Potentiation of human estrogen receptor alpha transcriptional activation through phosphorylation of serines 104 and 106 by the cyclin A-CDK2 complex. 1042 98

Cyclin-dependent kinases (CDKs) play an important role in the eukaryotic cell cycle progression. Cdc2 (CDK1) is expressed in late G(1)/S phase and required for G(2) to M phase transition in higher eukaryotes. The oncoproteins, SV40 large T antigen and adenovirus E1A, induce a 110-kDa protein which specifically recognizes the two inverted CCAAT motifs of the cdc2 promoter in cycling cells and plays an essential role in transactivation of the human cdc2 promoter. Since these CCAAT motifs also conform to the consensus binding sites for the ubiquitous heterotrimeric transcription factor, CBF/NF-Y, the role of CBF/NF-Y in the transactivation of the cdc2 promoter was examined in this study. Our results indicate that CBF/NF-Y and the 110-kDa protein interact with the CCAAT box motif to form a heteromeric complex. However, mutagenesis of the pentanucleotide CCAAT motif or in the presence of urea greater than 2.5 M, no heteromeric complex was formed. In contrast, the 110-kDa protein could still bind the mutant CCAAT motif or with the wild type motif in the presence of 2.5 M urea. Furthermore, E1A.12S induced the gene expression of all three subunits of CBF/NF-Y. Coexpression of E1A and a dominant negative mutant NF-YA subunit significantly reduced the E1A-mediated transactivation of the cdc2 promoter in a dose-dependent manner. These results support the conclusion that E1A protein mediates optimal transactivation of the human cdc2 promoter by inducing the expression and assembly of a heteromeric complex consisting of the 110-kDa protein and the CBF/NF-Y which interacts with the two CCAAT motifs of the cdc2 promoter.
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PMID:Transactivation of the human cdc2 promoter by adenovirus E1A. E1A induces the expression and assembly of a heteromeric complex consisting of the CCAAT box binding factor, CBF/NF-Y, and a 110-kDa DNA-binding protein. 1043 72


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