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Query: EC:2.7.11.22 (cdc2)
 8,319 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Neuronal cdc2-like kinase, nclk, is a heterodimer of cyclin dependent protein kinase 5, cdk5, and a 25 kDa subunit derived from a novel, neuron-specific, 35 kDa protein: p35. The characterization and regulation of nclk will be summarized in this minireview. The activity of nclk appears to be governed by highly complex regulatory mechanisms including protein-protein interaction, protein phosphorylation and isoforms. The histone H1 kinase activity of nclk is absolutely dependent of the interaction between the 25 kDa subunit and the catalytic subunit, cdk5. In addition, nclk interacts with other cellular proteins to form macromolecular complexes. The kinase activity of nclk is inhibited in vitro by the phosphorylation reactions of a weel-like protein tyrosine kinase and a protein serine/threonine kinase from bovine thymus. Northern blot analysis has revealed the existence of two populations of p35 mRNA of 2 and 4 kb. A novel cDNA encoding a p35 homologous protein has been obtained from a human hippocampus library.
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PMID:Regulatory properties of neuronal cdc2-like kinase. 856 47

We have isolated a novel member of putative serine/threonine kinase from a rat heart cDNA library using polymerase chain reaction methods. The novel kinase is transcribed as 2.6 kb mRNA encoding for a protein of 629 amino acids with the C-terminal non-catalytic portion. Amino acids analysis revealed that the N-terminal catalytic domain is 87% identical to the male-germ cell associated kinase (MAK), a cdc2-related serine/threonine kinase found to promote meiosis during spermatogenesis. Therefore, we designated this novel kinase as the MAK-related kinase (MRK). MRK protein, with a molecular weight of 66 kD, was shown to phosphorylate itself and the exogenous substrates, histone H1 and myelin basic protein. In addition, phosphoamino acid analysis confirmed the serine/threonine-specific protein kinase activity of MRK. Although MRK was ubiquitous in adult rat tissues, the expression of MRK protein in embryos was restricted primarily to embryonic myocardium during early organogenesis. This finding suggests that MRK may be a participant in cardiac development.
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PMID:Molecular cloning of a novel serine/threonine kinase, MRK, possibly involved in cardiac development. 857 Jan 68

Activation of p34cdc2 kinase is essential for entry into mitosis while subsequent deactivation and cyclin degradation are associated with exit. In Xenopus embryos, both of these phases are regulated by post-translation modifications and occur spontaneously on incubation of extracts prepared late in the first cell cycle. Even though high levels of calcium buffer were initially used to prepare these extracts, we found that free calcium levels in them remained in the observed physiological range (200-500 nM). Further addition of calcium buffers only slightly reduced free calcium levels, but inhibited histone H1 (cdc2A) kinase deactivation and cyclin degradation. Higher buffer concentrations slowed the kinase activation phase. Reducing the free buffer concentration by premixing with calcium reversed the effects of the buffer, indicating that the inhibitory effects arose from the calcium-chelating properties of the buffer rather than non-specific side effects. Furthermore, additions of calcium buffer at the end of the H1 kinase activation phase did not prevent deactivation. From these results, and the order of effectiveness of different calcium buffers in disrupting the H1 kinase cycle, we suggest that local transient increases in free calcium influence the rate of cdc2 kinase activation and are required to initiate the pathway leading to cyclin degradation and kinase inactivation in mitotic cell cycles.
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PMID:Calcium requirements during mitotic cdc2 kinase activation and cyclin degradation in Xenopus egg extracts. 858 67

To elucidate the role of protein kinase C in vascular smooth muscle cell proliferation, we examined the effects of phorbol 12-myristate 13-acetate (PMA) on G1 events in human arterial cells. About 15 h after G0 cells were stimulated with fetal bovine serum and basic fibroblast growth factor, [3H]thymidine incorporation started. PMA (10 nM) inhibited the incorporation over 90% when added earlier than 3 h after stimulation, but had no effect when added 12 h or later. PMA inhibited the phosphorylation of the retinoblastoma protein (pRb), which normally began at about 9 h. PMA did not inhibit the gene expression of Cdk2, Cdk3, Cdk4, Cdk5, and cyclins G, C, and D, all of which began at 0-3 h. However, PMA reduced the expression of cyclins E and A, which usually began at 3-9 h and about 15 h, respectively. PMA inhibited the histone H1 kinase activity of Cdk2, which increased from about 9 h, whereas PMA did not inhibit the pRb kinase activities of cyclin D-associated kinase(s) and Cdk4, detectable from 0-3 h. These results suggested that the PMA-induced inhibition of pRb phosphorylation is not mediated by suppressing cyclin D-associated kinase(s) including Cdk4, but involves the suppression of Cdk2 activity that results from the reduced expression of cyclins E and A.
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PMID:Phorbol ester inhibits the phosphorylation of the retinoblastoma protein without suppressing cyclin D-associated kinase in vascular smooth muscle cells. 862 31

Major transitions in the eukaryotic cell cycle are regulated by the cyclin-dependent protein kinases (CDK). In particular, the G2/M transition is initiated by the activity of a complex formed by a CDK of the Cdc2/Cdc28 family and B-type cyclins of the Cdc13/C1b family in the yeasts, Schizosaccharomyces pombe (Sp) and Saccharomyces cerevisiae (Sc). To study the molecular mechanisms that control the G2/M transition in the dimorphic pathogenic yeast, Candida albicans, we have cloned and characterized cDNAs corresponding to CDK1 and CYB1. The CDK1 cDNA encodes a 317-amino-acid (aa) protein that shares 76.8 and 62.3% identity with the Sc CDC28 and Sp cdc2 gene products, respectively. The CYB1 cDNA encodes a 493-aa protein that is 34.8, 34.4 and 35.5% identical to Sc C1b1 and C1b2, and to Sp Cdc13, respectively. Cyb1 contains characteristic mitotic destruction and cyclin boxes. The CDK1 and CYB1 cDNAs are functional homologues, as they are able to complement Sp cdc2 and cdc13 temperature-sensitive (ts) mutations, respectively, and their gene products interact in vivo in Sc to form an active histone H1 kinase.
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PMID:Candida albicans CDK1 and CYB1: cDNA homologues of the cdc2/CDC28 and cdc13/CLB1/CLB2 cell cycle control genes. 865 74

We have isolated a Leishmania mexicana homologue of the fission yeast suc1 gene using PCR with oligonucleotides designed to conserved regions of cdc2 kinase subunits (cks). The product of cks1 is a 12 kDa polypeptide, which has 70% identity with human p9cks1 and 44% identity with fission yeast p13suc1.p12cks1 was detected in the three life-cycle stages of L. mexicana by immunoblotting. Recombinant p12cks1 (p12cks1his) bound to agarose beads was used as a matrix to affinity-select histone H1 kinase complexes from Leishmania, yeast and bovine extracts. Immunoblotting showed that yeast and bovine cdc2 kinase bound to p12cks1his, thus demonstrating functional homology between L. mexicana p12cks1 and yeast p13suc1. Histone H1 kinase activity was found at a high level in the proliferative promastigote and amastigote forms of L. mexicana, but at a low level in the non-dividing metacyclic form. These activities are likely to be the same as the leishmanial p13suc1 binding kinase (SBCRK) described previously [Mottram, Kinnaird, Shiels, Tait and Barry (1993) J. Biol. Chem. 268, 21044-21051]. A distinct cdc2-related kinase, L. mexicana CRK1, was also found to associate with p12cks1his but affinity-depletion experiments showed that CRK1 was not responsible for the histone H1 kinase activity associating with p12cks1his in promastigote cell extracts. The finding that p12cks1 associates with at least two cdc2-related kinases, SBCRK and CRK1, is consistent with the presence of a large gene family of cdc2-related kinases in trypanosomatids, a situation thought to be more similar to higher eukaryotes than yeast.
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PMID:Leishmania mexicana p12cks1, a homologue of fission yeast p13suc1, associates with a stage-regulated histone H1 kinase. 867 Jan 59

Neuronal cdk5 can phosphorylate certain lys-ser-pro (KSP) motifs of neurofilaments and tau protein in the nervous system. We have immunoprecipitated the cdk5 from rat brain using a polyclonal antibody raised against the C-terminus of cdk5. The immunoprecipitate has phosphorylated a KSPXK peptide analog of NF-H, as well as histone H1 and a bacterially expressed rat NF-H protein. The kinase activity was inhibited by staurosporine, isopentanyladenine and olomoucine in a dose dependent manner. Kinetic studies indicated Ki values of 39 nM, 38 microM and 8 microM, respectively for staurosporine, isopentanyladenine and olomoucine. The inhibition by staurosporine was non-competitive with respect to phosphoryl acceptor acceptor substrates. Western blot analysis of the immunoprecipitate showed both cdk5 and p67 (Munc-18), a putative regulator molecule of the kinase. Addition of p67 fusion protein enhanced the kinase activity of the immunoprecipitate by 60% above the basal activity. P67 elevated Ki values for both staurosporine and olomoucine. The degree of inhibition at high concentrations of these inhibitors was unaltered by exogenous p67 indicating a lack of competitive interactions with p67. The high affinity of staurosporine for cdk5 suggests that cdk5 may be one of the targets for the neurotropic effect of staurosporine.
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PMID:Inhibition of neuronal cyclin-dependent kinase-5 by staurosporine and purine analogs is independent of activation by Munc-18. 872 73

The entry into mitosis is dependent on the activation of mitotic forms of cdc2 kinase. In many cell types, cyclin A-associated kinase activity peaks just prior to that of cyclin B, although the precise role of cyclin A-associated kinase in the entry into mitosis is still unclear. Previous work has suggested that while cyclin B is capable of triggering cyclin destruction in Xenopus cell-free systems, cyclin A-associated kinase is not able to support this function. Here we have expressed a full-length human cyclin A in Escherichia coli and purified the protein to homogeneity by virtue of an N-terminal histidine tag. We have found that when added to Xenopus cell-free extracts free of cyclin B and incapable of protein synthesis, the temporal pattern of cyclin A-associated cdc2 kinase activity showed distinct differences that were dependent on the concentration of cyclin A added. When cyclin A was added to a concentration that generated levels of cdc2 kinase activity capable of inducing nuclear envelope breakdown, the histone H1 kinase activity profile was bi-phasic, consisting of an activation phase followed by an inactivation phase. Inactivation was found to be due to cyclin destruction, which was prevented by mos protein. Cyclin destruction was followed by nuclear reassembly and an additional round of DNA replication, indicating that there is no protein synthesis requirement for DNA replication in this embryonic system. It has been suggested that the evolutionary recruitment of cyclin A into an S phase function may have necessitated the loss of an original mitotic ability to activate the cyclin destruction pathway. The results presented here indicate that cyclin A has not lost the ability to activate its own destruction and that cyclin A-mediated activation of the cyclin destruction pathway permitted destruction of cyclin B1 as well as cyclin A, indicating that there are not distinct cyclin A and cyclin B destruction pathways. Thus the ordered progression of the cell cycle requires the careful titration of cyclin. A concentration in order to avoid activation of the cyclin destruction pathway before sufficient active cyclin B/cdc2 kinase has accumulated.
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PMID:Activation of the Xenopus cyclin degradation machinery by full-length cyclin A. 874 54

We studied enzymatic activities in sea urchin egg extracts that phosphorylate myosin regulatory light chain (MRLC) from chicken gizzard smooth muscle. The activity in the presence of EGTA showed cell cycle-dependent changes similar to that of histone H1 kinase, namely, it peaked shortly before cleavage, while that in the presence of Ca2+ ions did not show significant change during division cycle. Phosphopeptide mapping revealed that both the sites phosphorylatable by smooth muscle myosin light chain kinase (MLCK sites) and the sites phosphorylatable by protein kinase C (PKC sites) were phosphorylated in the presence or absence of Ca2+ ions. By analyses using an inhibitor of cdc2 kinase, butyrolactone-I, and ion exchange column chromatography, at least three kinases were detected as kinases that phosphorylate MRLC in vitro. These kinases phosphorylated distinct sites on MRLC. The first one, which phosphorylated the PKC sites, was identified as cdc2 kinase. The second one phosphorylated the MLCK sites in the absence of Ca2+ ions. The third one phosphorylated unknown sites. Possible implication of these activities in regulation of cytokinesis is discussed.
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PMID:Cell cycle-dependent phosphorylation of smooth muscle myosin light chain in sea urchin egg extracts. 879 90

We have previously demonstrated that growth inhibition of untransformed intestinal epithelial cells by transforming growth factor beta1 (TGFbeta) and TGFbeta2 was associated with a rapid activation of both Ras and extracellular signal-regulated kinase 1 (Erk1) (Mulder, K. M., and Morris, S. L. (1992) J. Biol. Chem. 267, 5029-5031; Hartsough, M. T., and Mulder, K. M. (1995) J. Biol. Chem. 270, 7117-7124). In order to determine whether Ras was required for TGFbeta regulation of both Erk1 and downstream components associated with TGFbeta-mediated growth inhibition, the intestinal epithelial cell (IEC) line IEC 4-1 was transfected with a vector containing a dominant-negative mutant of Ras (RasN17) under the control of an inducible metallothionein promoter. Using two different RasN17-transfected clones treated with ZnCl2, we demonstrate here that induction of Ras expression by at least 4-fold completely abrogated the TGFbeta-mediated activation of Erk1. Moreover, the RasN17-mediated reversal of the TGFbeta effect on Erk1 was dependent upon the level of expression of the dominant-negative protein. ZnCl2 treatment of control cells transfected with the empty vector did not alter Ras expression or the activation of Erk1 by TGFbeta. In order to determine whether the activation of Ras by TGFbeta was required for the growth inhibitory effect of TGFbeta, we examined TGFbeta2 effects on Cdk2-associated histone H1 kinase activity, cyclin A protein expression levels, and DNA synthesis in two intestinal epithelial cell clones transfected with RasN17. In cells expressing RasN17, we observed a 50% reversal of the inhibition of Cdk2 activity, a 78% reversal of the down-regulation of cyclin A protein expression, and a 21% reversal of the inhibition of DNA synthesis by TGFbeta. Collectively, these results indicate that Ras activation is obligatory for TGFbeta-mediated activation of Erk1, whereas it is partially required for the growth inhibitory effect of TGFbeta.
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PMID:Altered transforming growth factor signaling in epithelial cells when ras activation is blocked. 879 98


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