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)

The cloning of the negative growth regulatory gene, p21Sdi1, has led to the convergence of the fields of cellular senescence, cell cycle regulation and tumor suppression. This gene was first cloned as an inhibitor of DNA synthesis that was overexpressed in terminally non-dividing senescent human fibroblasts (SD11) and later as a p53 transactivated gene (WAF1) and a Cdk-interacting protein (CIP1, p21) that inhibited cyclin-dependent kinase activity. To identify the active region(s) of p21Sdi1, cDNA constructs encoding various deleted forms of the protein were analyzed. Amino acids 22-71 were found to be the minimal region required for DNA synthesis inhibition. Amino acids 49-71 were involved in binding to Cdk2, and constructs deleted in this region expressed proteins that were unable to inhibit Cdk2 kinase activity in vitro. The latter stretch of amino acids shared sequence similarity with amino acids 60-76 of the p27Kip1 protein, another Cdk inhibitor. Point mutations made in p21Sdi1 in this region confirmed that amino acids common to both proteins were involved in DNA synthesis inhibition. Additionally, a chimeric protein, in which amino acids 49-65 of p21Sdi1 were substituted with amino acids 60-76 of p27Kip1, had almost the same DNA synthesis inhibitory activity as the wild-type protein. The results indicate that the region of sequence similarity between p21Sdi1 and p27Kip1 encodes an inhibitory motif characteristic of this family of Cdk inhibitors.
 ...
PMID:Identification of the active region of the DNA synthesis inhibitory gene p21Sdi1/CIP1/WAF1. 785 44

Core Binding Factor (CBF) is required for the development of definitive hematopoiesis, and the CBF oncoproteins AML1-ETO, TEL-AML1, and CBFbeta-SMMHC are commonly expressed in subsets of acute leukemia. CBFbeta-SMMHC slows the G1 to S cell cycle transition in hematopoietic cells, but the mechanism of this effect is uncertain. We have sought to determine whether inhibition of CBF-mediated trans-activation is sufficient to slow proliferation. We demonstrate that activation of KRAB-AML1-ER, a protein containing the AML1 DNA-binding domain, the KRAB repression domain, and the Estrogen receptor ligand binding domain, also slows G1, if its DNA-binding domain is intact. Also, exogenous AML1 overcame CBFbeta-SMMHC-induced inhibition of proliferation. Representational difference analysis (RDA) identified cdk4 RNA expression as an early target of KRAB-AML1 activation. Inhibition of CBF activities by KRAB-AML1-ER or CBFbeta-SMMHC rapidly reduced endogenous cdk4 mRNA levels, even in cells proliferating at or near control rates as a result of exogenous cdk4 expression. Over-expression of cdk4, especially a variant which cannot bind p16INK4a, overcame cell cycle inhibition resulting from activation of KRAB-AML1-ER, although cdk4 did not accelerate proliferation when expressed alone. These findings indicate that mutations which alter the expression of G1 regulatory proteins can overcome inhibition of proliferation by CBF oncoproteins. Oncogene (2000).
 ...
PMID:Exogenous cdk4 overcomes reduced cdk4 RNA and inhibition of G1 progression in hematopoietic cells expressing a dominant-negative CBF - a model for overcoming inhibition of proliferation by CBF oncoproteins. 1085 Oct 69

Gene expression of the plasminogen activation system is cell-cycle dependent. Previously, we showed that ectopic expression of E2F1 repressed the plasminogen activator inhibitor type 1 (PAI-1) promoter in a manner dependent on the presence of DNA-binding and transactivation domains of E2F1 but independent of binding to pocket-binding proteins, suggesting a novel mechanism for E2F-mediated negative gene regulation [Koziczak, M., Krek, W. & Nagamine, Y. (2000) Mol. Cell. Biol. 20, 2014-2022]. However, it remains to be seen whether endogenous E2F can exert a similar effect. We report here that down-regulation of PAI-1 gene expression correlates with an increase in endogenous E2F activity. When cells were treated with a cdk2/4-specific inhibitor, which maintains E2F in an inactive state, the decline of serum-induced PAI-1 mRNA levels was suppressed. In mutant U2OS cells expressing a temperature-sensitive retinoblastoma protein (pRB), a shift to a permissive temperature induced PAI-1 mRNA expression. In U2OS cells stably expressing an E2F1-estrogen receptor chimeric protein that could be activated by tamoxifen, PAI-1 gene transcription was markedly reduced by tamoxifen even in the presence of cycloheximide. These results all indicate that endogenous E2F can directly repress the PAI-1 gene. DNase I hypersensitive-site analysis of the PAI-1 promoter suggested the involvement of conformation changes in chromatin structure of the PAI-1 promoter. 5' deletion analysis of the PAI-1 promoter showed that multiple sites were responsible for the E2F negative regulation, some of which were promoter dependent. Interestingly, one of these sites is a p53-binding element.
 ...
PMID:E2F1-mediated transcriptional inhibition of the plasminogen activator inhibitor type 1 gene. 1155 66

Cdc25 is a dual-specificity phosphatase that catalyzes the activation of the cyclin-dependent kinases (Cdk/cyclins), thus triggering initiation and progression of successive phases of the cell cycle. In our efforts to elucidate the interaction between Cdc25B and the natural substrate, bis-phosphorylated Cdk2/CycA (Cdk2-pTpY/CycA), we have previously found that the 17 residues of the C-terminal tail mediate a factor of 10 in substrate recognition. In the studies reported here, we localize the majority of this interaction using site-directed mutagenesis to two arginine residues (Arg556 and Arg562) located within this C-terminal region. We also show that the catalytic domain of Cdc25C, which differs most significantly from Cdc25B in this tail region, has a 100-fold lower activity toward Cdk2-pTpY/CycA. We further demonstrate that the proper presentation of the C-terminal tail of Cdc25B can be achieved in a "gain-of-function" chimeric protein consisting of the C-terminal tail of Cdc25B fused onto the catalytic core of Cdc25C. The >10-fold increase in activity seen only in the chimeric protein containing the two critical arginine residues demonstrates that the modular C-terminal tail of Cdc25B is the basis for most of the catalytic advantage of Cdc25B versus Cdc25C toward the Cdk2-pTpY/CycA substrate.
 ...
PMID:The C-terminal tail of the dual-specificity Cdc25B phosphatase mediates modular substrate recognition. 1171 73

Earlier studies have shown that translation elongation factor 1delta (EF-1delta) is hyperphosphorylated in various mammalian cells infected with representative alpha-, beta-, and gammaherpesviruses and that the modification is mediated by conserved viral protein kinases encoded by herpesviruses, including UL13 of herpes simplex virus type 1 (HSV-1), UL97 of human cytomegalovirus, and BGLF4 of Epstein-Barr virus (EBV). In the present study, we attempted to identify the site in EF-1delta associated with the hyperphosphorylation by the herpesvirus protein kinases. Our results are as follows: (i) not only in infected cells but also in uninfected cells, replacement of the serine residue at position 133 (Ser-133) of EF-1delta by alanine precluded the posttranslational processing of EF-1delta, which corresponds to the hyperphosphorylation. (ii) A purified chimeric protein consisting of maltose binding protein (MBP) fused to a domain of EF-1delta containing Ser-133 (MBP-EFWt) is specifically phosphorylated in in vitro kinase assays by purified recombinant UL13 fused to glutathione S-transferase (GST) expressed in the baculovirus system. In contrast, the level of phosphorylation by the recombinant UL13 of MBP-EFWt carrying an alanine replacement of Ser-133 (MBP-EFS133A) was greatly impaired. (iii) MBP-EFWt is also specifically phosphorylated in vitro by purified recombinant BGLF4 fused to GST expressed in the baculovirus system, and the level of phosphorylation of MBP-EFS133A by the recombinant BGLF4 was greatly reduced. (iv) The sequence flanking Ser-133 of EF-1delta completely matches the consensus phosphorylation site for a cellular protein kinase, cdc2, and in vitro kinase assays revealed that purified cdc2 phosphorylates Ser-133 of EF-1delta. (v) As observed with EF-1delta, the casein kinase II beta subunit (CKIIbeta) was specifically phosphorylated by UL13 in vitro, while the level of phosphorylation of CKIIbeta by UL13 was greatly diminished when a serine residue at position 209, which has been reported to be phosphorylated by cdc2, was replaced with alanine. These results indicate that the conserved protein kinases encoded by herpesviruses and a cellular protein kinase, cdc2, have the ability to target the same amino acid residues for phosphorylation. Our results raise the possibility that the viral protein kinases mimic cdc2 in infected cells.
 ...
PMID:Conserved protein kinases encoded by herpesviruses and cellular protein kinase cdc2 target the same phosphorylation site in eukaryotic elongation factor 1delta. 1255 73

Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) is a phosphoprotein suggested to play important roles in EBV-induced immortalization. Earlier studies have shown that the major site of phosphorylation of EBNA-LP by cellular kinase(s) is a serine residue at position 35 (Ser-35) and that the phosphorylation of Ser-35 is critical for regulation of the coactivator function of EBNA-LP (Yokoyama et al., J Virol 75, 5119-5128, 2001). In the present study, we have attempted to identify protein kinase(s) responsible for the phosphorylation of EBNA-LP at Ser-35. A purified chimeric protein consisting of glutathione S-transferase (GST) fused to a domain of EBNA-LP containing Ser-35 was found to be specifically phosphorylated by purified cdc2 in vitro, while GST fused to a mutated domain of EBNA-LP in which Ser-35 was replaced with alanine was not. In addition, overexpression of cdc2 in mammalian cells caused a significant increase in the phosphorylation of EBNA-LP, while this increased phosphorylation was eliminated if Ser-35 of EBNA-LP was replaced with alanine. These results indicate that the cellular protein kinase cdc2 mediates the phosphorylation of EBNA-LP at Ser-35. Recently, we reported that cdc2 and conserved protein kinases encoded by herpesviruses phosphorylate the same amino acid residue of target proteins (Kawaguchi et al., J Virol 77, 2359-2368, 2003). Consistent with this, the EBV-encoded conserved protein kinase BGLF4 specifically mediated the phosphorylation of EBNA-LP at Ser-35. These results indicate that the coactivator function of EBNA-LP can be regulated by the activity of these cellular and viral protein kinases.
 ...
PMID:Identification of protein kinases responsible for phosphorylation of Epstein-Barr virus nuclear antigen leader protein at serine-35, which regulates its coactivator function. 1464 19

Cyclin-dependent kinases (cdks) represent potentially promising molecular targets for cancer therapeutic strategies. To evaluate the antitumor activity of selective cyclin/cdk inhibition, we constructed a chimeric protein composed of a F-box protein (TrCP) fused to a peptide comprising the cyclin/cdk2 binding motif in p21-like cdk inhibitors (TrCP-LFG). We now demonstrate that endogenous cyclin A and its binding substrate, cdk2, can be tethered to beta-TrCP, ubiquitinated, and effectively degraded. Degradation of cdk2 and cyclin A together, but not cdk2 alone, results in massive tumor cell apoptosis in vitro and in vivo in a proteasome-dependent manner with no toxicity to normal tissue. These data demonstrate that cyclin A and/or the cyclin A/cdk2 complex is a promising anticancer target with a high therapeutic index.
 ...
PMID:Proteasome-mediated destruction of the cyclin a/cyclin-dependent kinase 2 complex suppresses tumor cell growth in vitro and in vivo. 1517 7

MEF is an ETS-related transcription factor with strong transcriptional activating activity that affects hematopoietic stem cell behavior and is required for normal NK cell and NK T-cell development. The MEF (also known as ELF4) gene is repressed by several leukemia-associated fusion transcription factor proteins (PML-retinoic acid receptor alpha and AML1-ETO), but it is also activated by retroviral insertion in several cancer models. We have previously shown that cyclin A-dependent phosphorylation of MEF largely restricts its activity to the G(1) phase of the cell cycle; we now show that MEF is a short-lived protein whose expression level also peaks during late G(1) phase. Mutagenesis studies show that the rapid turnover of MEF in S phase is dependent on the specific phosphorylation of threonine 643 and serine 648 at the C terminus of MEF by cdk2 and on the Skp1/Cul1/F-box (SCF) E3 ubiquitin ligase complex SCF(Skp2), which targets MEF for ubiquitination and proteolysis. Overexpression of MEF drives cells through the G(1)/S transition, thereby promoting cell proliferation. The tight regulation of MEF levels during the cell cycle contributes to its effects on regulating cell cycle entry and cell proliferation.
 ...
PMID:The ETS protein MEF is regulated by phosphorylation-dependent proteolysis via the protein-ubiquitin ligase SCFSkp2. 1658 86

The fusion gene AML1-ETO initially dysregulates various cell cycle molecules in t(8;21) acute myeloid leukemia. Aurora kinases have shown great promise in treating tumors. However, the efficacy of Aurora kinase (AURK) A and B inhibition in t(8;21) AML remains unclear. We found that AURK-A inhibitor Alisertib and AURK-B inhibitor Barasertib strongly inhibited the growth and proliferation of t(8;21) AML cells. The quantity and size of cell colonies were markedly decreased after a 14-d drug exposure. The cell cycle distribution was blocked at the G2/M phase in both dose- and time-dependent manner. The expression of p53 family and cdc2-p34 significantly changed as well. Notably, we found that t(8;21) AML cells are more sensitive to Aurora B inhibition. In each set of experiments, Barasertib took less time or a lower concentration to achieve similar efficacy. Taken together, our data highlighted the potential role of Aurora kinases as promising cell cycle targets for the treatment of t(8;21) AML and hereby provided a theoretical basis to guide relevant clinical trials.
 ...
PMID:Selective inhibition of Aurora A and B kinases effectively induces cell cycle arrest in t(8;21) acute myeloid leukemia. 3120 77