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 cloned, sequenced and disrupted the checkpoint genes RAD17, RAD24 and MEC3 of Saccharomyces cerevisiae. Mec3p shows no strong similarity to other proteins currently in the database. Rad17p is similar to Rec1 from Ustilago maydis, a 3' to 5' DNA exonuclease/checkpoint protein, and the checkpoint protein Rad1p from Schizosaccharomyces pombe (as we previously reported). Rad24p shows sequence similarity to replication factor C (RFC) subunits, and the S. pombe Rad17p checkpoint protein, suggesting it has a role in DNA replication and/or repair. This hypothesis is supported by our genetic experiments which show that overexpression of RAD24 strongly reduces the growth rate of yeast strains that are defective in the DNA replication/repair proteins Rfc1p (cdc44), DNA pol alpha (cdc17) and DNA pol delta (cdc2) but has much weaker effects on cdc6, cdc9, cdc15 and CDC4 strains. The idea that RAD24 overexpression induces DNA damage, perhaps by interfering with replication/repair complexes, is further supported by our observation that RAD24 overexpression increases mitotic chromosome recombination in CDC4 strains. Although RAD17, RAD24 and MEC3 are not required for cell cycle arrest when S phase is inhibited by hydroxyurea (HU), they do contribute to the viability of yeast cells grown in the presence of HU, possibly because they are required for the repair of HU-induced DNA damage. In addition, all three are required for the rapid death of cdc13 rad9 mutants. All our data are consistent with models in which RAD17, RAD24 and MEC3 are coordinately required for the activity of one or more DNA repair pathways that link DNA damage to cell cycle arrest.
Mol Gen Genet 1997 Nov
PMID:G2/M checkpoint genes of Saccharomyces cerevisiae: further evidence for roles in DNA replication and/or repair. 943 89

It is now well established that progression through the eukaryotic cell cycle is controlled by oscillations in the activity of cyclin- dependent kinases (CDKs). In many cases, however, the physiological substrate(s) of CDKs are unknown. The Saccharomyces cerevisiae PHO5 gene encodes a secreted acid phosphatase which is induced in response to phosphate starvation. The PHO5 gene is activated by the Pho4p transcription factor, which itself is negatively regulated through phosphorylation by the products of PHO80 and PHO85. Pho80p and Pho85p are homologous to cyclins and CDKs, respectively, and the Pho80p/Pho85p heterodimer satisfies the biochemical definition of a cyclin/CDK. In the present study, several reporter genes were expressed in S. cerevisiae from promoters which are activated by the transcription factor Pho4p, thereby generating yeast strains which exhibit quantifiable phenotypes that reflect the activity of a specific cyclin/CDK. Positive genetic selections for inhibition of cyclin/CDK function were characterized using the E. coli neo and yeast LEU2 genes. Chromosomal disruptions of the yeast PHO80 and PHO85 genes were constructed and conditions for complementation by plasmid-borne genes were defined. Complementation is achieved at very low levels of expression of both Pho80p and Pho85p. High-level expression of Pho80p results in aberrant PHO5 promoter regulation, characterized by failure to derepress in low-phosphate medium. Genes encoding hybrid CDKs in which regions of Pho85p were replaced with the homologous region of human Cdk2 were constructed, and tested for function in S. cerevisiae by complementation of the pho85 chromosomal gene disruption. Hybrid proteins in which more than two-thirds of the molecule were derived from human Cdk2 retained Pho85p function with respect to high-phosphate repression of the PHO5 promoter. The hybrid proteins require the PHO80 gene product for this function. A hybrid human-yeast CDK in which a single amino acid is deleted, within a nonapeptide sequence which is perfectly conserved in Pho85p and human Cdk2, retains full function. These results demonstrate that, within the context of the conserved structure of CDKs, considerable primary sequence variability can be introduced without loss of the cyclin-dependent function of the CDK.
Mol Gen Genet 1998 Oct
PMID:Function of hybrid human-yeast cyclin-dependent kinases in Saccharomyces cerevisiae. 982 36

Human herpesvirus-8 (HHV-8), also known as Kaposi's sarcoma-associated herpesvirus, encodes a protein, referred to as HHV8-Vcyc, with sequence similarity to human G1 cyclins, in particular of the D type. HHV8-Vcyc is expressed in Kaposi's sarcoma and functional analysis suggests that it can activate cyclin-dependent kinases (cdk) and thereby trigger inactivation of the retinoblastoma protein (pRb), indicating that HHV8-Vcyc may contribute to the oncogenic potential of HHV-8. We show here that HHV8-Vcyc can activate transcription of the human cyclin A gene in quiescent cells, a property shared with known transforming oncogenes. Transcriptional activation by HHV8-Vcyc depends on an E2F-binding site in the cyclin A promoter, and cdk6 kinase activity is required. The ability of HHV8-Vcyc to activate cyclin A gene expression is shared by D-type cyclins and cyclin E. Unlike D-type cyclins, HHV8-Vcyc is unable to trigger phosphorylation of the pRb-related protein p107 and fails to induce dissociation of p107 from E2F. Unlike cyclin E, HHV8-Vcyc fails to interact physically with E2F complexes on the cyclin A promoter. These results provide additional evidence for the notion that the HHV-8-encoded cyclin differs in several properties from cellular G1 cyclins.
J Gen Virol 1999 Mar
PMID:Activation of cyclin A gene expression by the cyclin encoded by human herpesvirus-8. 1009 92

In Schizosaccharomyces pombe, wee1 encodes a tyrosine kinase that inhibits entry into mitosis by phophorylating Cdc2, the universal cyclin-dependent kinase (Cdk) that regulates the G2/M transition in all eukaryotic cells. A search for suppressors of the G2 arrest caused by overexpression of weel led to the isolation of a new allele of swo1 (named swo1-w1), the gene coding for chaperone Hsp90, which is required to stabilise Weel. The swo1-w1 allele carries a glycine to aspartic acid substitution at amino acid 155 that results in a partial loss of Hsp90 function. Cells bearing the swo1-w1 mutation in combination with the point mutation cdc2-33 or cdc2-M26 showed severe mitotic defects. Genetic interactions were not observed in combination with point mutations in other cdc genes, suggesting that Cdc2 specifically interacts with Hsp90. This synthetic lethal swo1-w1 cdc2-33 (or cdc2-M26) strain had normal levels of Cdc2 protein and histone H1 phosphorylation activity, indicating that Hsp90 is required to enable Cdc2 to interact with its mitotic substrates or regulators, rather than for its proper folding or stabilisation. In a wild-type background, swo1-w1 mutant cells were sensitive to temperature as well as to other stress agents, such as KCI, ethanol and formamide. Under these stressful growth conditions, the swo1-w1 cells displayed anaphase B arrest and aberrant septation patterns, indicating that a subset of proteins involved in mitosis and cytokinesis is highly dependent on chaperone Hsp90 for function.
Mol Gen Genet 1999 Mar
PMID:Genetic interactions between Hsp90 and the Cdc2 mitotic machinery in the fission yeast Schizosaccharomyces pombe. 1010 58

The kinase Dbf4p/Cdc7p is required for the G1/S phase transition during the cell cycle and plays a direct role in the activation of individual origins of replication in Saccharomyces cerevisiae. Here, we report the identification and characterization of mouse and human cDNAs whose products are related in sequence to Saccharomyces cerevisiae DBF4 cDNA. Both mammalian Dbf4 proteins contain a putative site for phosphorylation by CDK, PEST protease cleavage sites, nuclear localization signals and a short-looped zinc finger-like domain. Transcription of MmDBF4 is suppressed in mouse NIH3T3 fibroblasts made quiescent by serum starvation. Upon replenishment of the medium, transcript levels increase during progression through G1, peaking as cells enter S phase. MmDbf4p interacts physically with Cdc7p and Mcm2p in vivo. Using fluorescence in situ hybridization (FISH), the human DBF4 gene was localized to chromosome 7 (q21.3), whereas FISH mapped the murine counterpart to band A2 on chromosome 5. The results of chromosome mapping indicate that in both mouse and human the gene is present as a single copy. The structural conservation between Dbf4-related proteins suggests that these proteins play a key role in the regulation of DNA replication during the cell cycle in all eukaryotes.
Mol Gen Genet 1999 Sep
PMID:Identification, characterization and chromosomal localization of the cognate human and murine DBF4 genes. 1051 17

Cyclins are known to activate cyclin-dependent protein kinases, which are essential for cell cycle progression in eukaryotes. We isolated full-length cDNAs encoding rice mitotic cyclins named CycA1; os; 1 and CycB2;os;1, which are related to A- and B-type cyclins, respectively, from animals. To characterize the function of these mitotic cyclins, as well as that of another B-type cyclin, CycB2;os;2, each cDNA was introduced into yeast cells. When cDNAs encoding CycA1;os;1, CycB2;os; or CycB2;os;2 were overexpressed in the yeast mutant DLI, which is deficient in G1 cyclins, the mutant phenotype was rescued, indicating that these mitotic cyclins are functional in yeast cells. When the cDNA encoding CycB2;os;1 was expressed in the wild-type yeast strain, the cells lost the ability to grow, whereas the expression of either cycA1;os: 1 or cycB2;os;2 did not inhibit growth. In situ hybridization of these mitotic cyclin genes with rice root apices and counterstaining of chromosomes with a DNA-specific dye revealed that cycA1;os;1 is expressed from the G2 phase to the early M phase, while transcripts of cycB2:os;1 and cycB2;os;2 accumulated until the end of mitosis. Our results indicate that these B2-type cyclins may be involved in the control of mitosis, in combination with a G2/M-phase CDK.
Mol Gen Genet 1999 Sep
PMID:Molecular characterization of mitotic cyclins in rice plants. 1051 18

The yeast transcription factor Ace2p regulates expression of the chitinase gene CTS1 in a cell cycle-dependent manner. Nuclear localisation of Ace2p is restricted to late M and early G phases of the mitotic cell cycle. We show here that this nuclear localisation is directly associated with regulation of CTS1 expression. Using a version of Ace2p tagged with a c-myc epitope, we show that the protein is excluded from the nucleus of cells during most phases of the mitotic cell cycle. A mutant derivative in which one threonine and two serine residues, which are candidate phosphorylation sites, were replaced by alanine (to mimic constitutive dephosphorylation) is localised in the nucleus throughout the cell cycle. The mechanism of localisation of Ace2p therefore involves regulation of its phosphorylation state, and closely resembles that used by the homologous transcription factor Swi5p. The wild-type Ace2 protein associates with Cdc28p in vivo, suggesting this may be the kinase that mediates the phosphorylation event. The stability of the protein is greatly reduced in a mutant that is constitutively localised to the nucleus, but is restored in a deletion derivative which remains in the cytoplasm. Ace2p is therefore controlled throughout the cell cycle at three levels: transcription, nuclear localisation, and proteolysis.
Mol Gen Genet 1999 Sep
PMID:Regulated nuclear localisation of the yeast transcription factor Ace2p controls expression of chitinase (CTS1) in Saccharomyces cerevisiae. 1051 23

The Saccharomyces cerevisiae genes PHO80 and PHO85 encode, respectively, a cyclin and cyclin-dependent kinase, which negatively regulate PHO5 gene transcription by phosphorylating the transcription activator Pho4p. Cyclin-dependent kinases (CDKs) are highly conserved proteins, both within and between species. It was previously demonstrated, using reporter genes activated in yeast by Pho4p, that hybrid proteins in which over two-thirds of Pho85p were replaced with the homologous region from human Cdk2 retained the function of native Pho85p with respect to promoter repression. In the present study, various truncated forms of the hybrid human-yeast CDKs were tested for function. Surprisingly, truncations in which significant portions of the C-terminal region of the 291-residue hybrid CDK were deleted retained activity. Genes encoding human Cdk2 proteins which terminated after amino acids 151, 140, 130, 120 and 90 each complement a chromosomal pho85 gene disruption in which the HIS3 gene is inserted at codon 49. Truncated Cdk2 proteins containing less than 60 amino acids failed to complement the pho85::HIS3 gene disruption. Although the functional C-terminal truncations disrupt the ATP-binding and active sites of Cdk2, reporter gene repression mediated by these truncated proteins is apparently due to phosphorylation of Pho4p, since a gene in which the essential lysine codon at position 33 was converted to an arginine codon does not complement the chromosomal gene disruption. The human Cdk2 truncations were demonstrated to function through intergenic complementation. The intact Cdk2-Pho85 hybrid CDK complemented the pho85 mutation in yeast strains in which the entire PHO85 coding region was deleted from chromosome XVI. The C-terminal Cdk2 truncations, however, were non-functional in these strains and thus dependent for activity on the pho85 coding region which remained in the mutant pho85::HIS3 chromosomal locus. These genetic results are consistent with a model involving protein fragment complementation in which the active site of the CDK is bisected.
Mol Gen Genet 2000 Mar
PMID:Intergenic complementation truncation mutants of cyclin-dependent kinase. 1077 40

Hyperactivation of Cdc2 in fission yeast causes cells to undergo a lethal premature mitosis, a phenomenon called mitotic catastrophe. This phenotype is observed in cdc2-3w wee1-50 cells at high temperature and is suppressed by a single recessive mutant, mcs3-12. Mcs3 acts independently of the Wee1 kinase and Cdc25 phosphatase, two major regulators of Cdc2. We have isolated multicopy suppressors of the cell cycle arrest phenotype of mcs3-12 wee1-50 cdc25-22 cells, but did not identify the mcs3 gene itself. Instead several known mitotic regulators were isolated, including the Cdc25 phosphatase, Wis2 cyclophilin, Cek1 kinase, and an Hsp90 homologue, Swo1. We also isolated clones encoding non-functional, truncated forms of the Wee1 kinase and Dis2 type 1 phosphatase. In addition we identified a multicopy suppressor that encodes a structural homologue of the budding yeast SPO12 gene. We find that overexpression of fission yeast spo12 not only suppresses the phenotype of the mcs3-12 wee1-50 cdc25-22 strain, but also that of a win1-1 wee1-50 cdc25-22 strain at high temperature, indicating that the function of spo12 is not directly related to mcs3. We show that spo12 mRNA is periodically expressed during the fission yeast cell cycle, peaking at the G2/M transition coincidently with cdc15. Deletion of spo12, however, has no overt effect on either the mitotic or meiotic cell cycles, except when the function of the major B type cyclin, Cdc13, is compromised.
Mol Gen Genet 2000 Oct
PMID:spo12 is a multicopy suppressor of mcs3 that is periodically expressed in fission yeast mitosis. 1108 71

The cyclin-dependent kinase inhibitor p27KIP1 plays a key role in controlling cell proliferation. Here we show that p27KIP1 is commonly down-regulated in B-cells immortalized by Epstein-Barr virus (EBV) (lymphoblastoid cell lines, LCLs). The significance of this event for the immortal phenotype of LCLs is implied by a requirement for active cdk2-containing complexes for continued proliferation, and by the ability of the residual p27KIP1 to associate with cdk2. The mechanism of p27KIP1 attenuation is post-translational, but inhibitor studies reveal that the mechanism does not rely heavily on the proteasome. Instead we find that LCLs contain an activity that cleaves a caspase recognition site present in p27KIP1 (DPSD139). The activity is not associated with apoptosis and closely resembles a proliferation-associated caspase activity we previously described in the EBV-negative B-lymphoma-derived cell line BJAB. Importantly, proliferating LCLs contain a p27KIP1 product that is consistent with cleavage at this site. Inhibition of caspase(s) in vivo modulates p27KIP1 expression and strongly inhibits proliferation of IB4 cells. This inhibitor profile is identical to that displayed by the DPSD-directed caspase present in BJAB cells, suggesting that the caspase may fulfil a general role in controlling p27KIP1 expression in immortal lymphoid cell lines. Thus, apoptosis-independent cleavage appears to contribute to the maintenance of the low basal levels of p27KIP1 in B-cells immortalized by EBV.
J Gen Virol 2001 Dec
PMID:Regulation of p27KIP1 in Epstein-Barr virus-immortalized lymphoblastoid cell lines involves non-apoptotic caspase cleavage. 1171 84


<< Previous 1 2 3 4 Next >>