Gene/Protein Disease Symptom Drug Enzyme Compound
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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)

Progression of BY-2 tobacco cells through the cell cycle was followed after treatments with ultra violet (UV) and salicylic acid (SA) used as a potent inhibitor of the octadecanoid pathway which can mediate response to UV irradiation. Cells in S phase were more sensitive than G0/G1 or G2 cells to UV irradiation. Although SA efficiently blocked cells in G0/G1 or G2, it did not block S phase synchronized cells. UV and SA applied simultaneously to cells in G0/G1 delayed the cell cycle progression more than each one separately. Therefore UV irradiation and SA act as agonists to arrest BY-2 cells at cell cycle entry. To further investigate the signalling pathway mediating UV response, we complemented a UV-sensitive Escherichia coli strain with a Nicotiana xanthi cDNA expression library. A cDNA (arcA3) whose coding sequence is identical to the 2,4-D induced arcA cDNA cloned by Ishida et al. (1993) was isolated. We show that arcA3 transcription is induced at cell cycle entry but not directly by the 2,4-D treatment. Moreover, arcA3 transcription is induced prior to the restriction point as shown with the CDK inhibitor roscovitine. The arcA3 transcription level is increased by UV irradiation but prevented by SA. Indeed, addition of SA prior to UV irradiation blocks the induction of arcA3 transcription. This suggests that arcA3 gene is modulated in both UV and SA responses, the SA effect preceding the UV step. Since arcA3 is 67% similar to RACK1 (functional homology), a rat intracellular receptor for protein kinase C, and possesses identical PKC fixation motifs, it is hypothesised that the arcA3 gene is involved in UV and SA cell cycle arrest.
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PMID:Is arcA3 a possible mediator in the signal transduction pathway during agonist cell cycle arrest by salicylic acid and UV irradiation? 1008 53

Expression of full-length p16(INK4a) blocks alphavbeta3 integrin-dependent cell spreading on vitronectin but not collagen IV. Similarly, G1-associated cell cycle kinases (CDK) inhibitory (CKI) synthetic peptides derived from p16(INK4a), p18(INK4c) and p21(Cip1/Waf1), which can be delivered directly into cells from the tissue culture medium, do not affect non-alphavbeta3-dependent spreading on collagen IV, laminin and fibronectin at concentrations that inhibit cell cycle progression in late G1. The alphavbeta3 heterodimer remains intact after CKI peptide treatment but is immediately dissociated from the focal adhesion contacts. Treatment with phorbol 12-myristate 13-acetate (PMA) allows alphavbeta3 to locate to the focal adhesion contacts and the cells to spread on vitronectin in the presence of CKI peptides. The cdk6 protein is found to suppress p16(INK4a)-mediated inhibition of spreading and is also shown to localize to the ruffling edge of spreading cells, indicating a function for cdk6 in controlling matrix-dependent cell spreading. These results demonstrate a novel G1 CDK-associated integrin regulatory pathway that acts upstream of alphavbeta3-dependent activation of PKC as well as a novel function for the p16(INK4a) tumour suppressor protein in regulating matrix-dependent cell migration.
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PMID:The p16(INK4a) tumour suppressor protein inhibits alphavbeta3 integrin-mediated cell spreading on vitronectin by blocking PKC-dependent localization of alphavbeta3 to focal contacts. 1020 65

The myristoylated alanine-rich C-kinase substrate (MARCKS) purified from brain was recently characterized as a proline-directed kinase(s) substrate in vivo [Taniguchi, Manenti, Suzuki and Titani (1994) J. Biol. Chem. 269, 18299-18302]. Here we have investigated the phosphorylation of MARCKS by various cyclin-dependent kinases (Cdks) in vitro. We established that Cdk2, Cdk4 and, to a smaller extent, Cdk1 that have been immunoprecipitated from cellular extracts phosphorylate MARCKS. Comparison of MARCKS phosphorylation by protein kinase C (PKC) and by the purified cyclin E-Cdk2 complex suggested that two residues were phosphorylated by Cdk2 under these conditions. To identify these sites, Cdk2-phosphorylated MARCKS was digested with lysyl endoprotease and analysed by electrospray MS. Comparison with the digests obtained from the unphosphorylated protein demonstrated that two peptides, Gly12-Lys30 and Ala138-Lys152, were phosphorylated by cyclin E-Cdk2. The identity of these peptides was confirmed by automatic Edman degradation. On the basis of the consensus phosphorylation sequence described for Cdk2, and on MS/MS analysis of the Ala138-Lys152 peptide, we concluded that Ser27, one of the phosphorylation sites identified in vivo, and Thr150 were the Cdk2 targets in vitro. None of the other sites described in vivo were phosphorylated in these conditions. Interestingly, a preliminary phosphorylation of MARCKS by PKC improved the initial rate of phosphorylation by Cdk2 without modifying the number of sites concerned. In contrast, phosphorylation of MARCKS by Cdk2 did not significantly affect further phosphorylation by PKC.
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PMID:Phosphorylation of the myristoylated protein kinase C substrate MARCKS by the cyclin E-cyclin-dependent kinase 2 complex in vitro. 1035 64

Signal transduction mediated by the single yeast isozyme of protein kinase C (Pkc1p) is essential for the maintenance of cellular integrity in this model eukaryote. The past few years have seen a dramatic increase in our knowledge of the upstream regulatory factors that modulate Pkc1p activity (e.g. Tor2p, Rom1p, Rom2p, Rho1p, Slg1p, Mid2p) and of the downstream targets of the MAP kinase cascade triggered by it (e.g. Rlm1p, SBF complex). The picture that has emerged connects this pathway to a variety of other cellular processes, such as cell cycle progression (Cdc28p, Swi4p), mating (Ste20p), nutrient sensing (Ira1p), calcium homeostasis (calcineurin, Mid2p, Fks2p) and the structural dynamics of the cytoskeleton (Spa1p, Bni1p).
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PMID:The protein kinase C-mediated MAP kinase pathway involved in the maintenance of cellular integrity in Saccharomyces cerevisiae. 1036 Dec 72

We have combined a rapid cytoplasmic sampling technique with capillary electrophoresis to measure the activation of protein kinase C (PKC) in a small region (approximately 60 microm) of a Xenopus oocyte. The phosphorylation of a fluorescent PKC substrate was measured following addition of a pharmacological or physiological stimulus to an oocyte. When substrates for cdc2 kinase (cdc2K), PKC, and protein kinase A (PKA) were comicroinjected into an oocyte, all three substrates could be identified on the electropherogram after cytoplasmic sampling. With this new method, it should be possible to measure simultaneously the activation of multiple different kinases in a single cell, enabling the quantitative dissection of signal transduction pathways.
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PMID:Localized measurement of kinase activation in oocytes of Xenopus laevis. 1042 39

Although widely used as an operational marker of proliferation, the cell cycle-regulated Ki67 protein is of unknown function. pKi67 is found predominantly in the nucleolus in cycling interphase cells and moves to become perichromosomal during mitosis. We have performed a detailed immunochemical analysis of pKi67 in HeLa cells and report the existence of a novel hyperphosphorylated form in mitosis. Two isoforms can be identified on immunoblots as a consequence of the previously described alternative splicing. In extracts from mitotic cells both these isoforms have considerably reduced mobility. Treatment with phosphatase converts the mitotic form to the interphase form. Immunoprecipitated pKi67 can be phosphorylated in vitro both by cdc2/cyclin B and by protein kinase C, and treatment by PKC leads to the full mobility shift. Treatment of nocodazole-arrested mitotic HeLa cells with staurosporine causes a dephosphorylation of pKi67 to the interphase state and a concomitant change in the localization of pKi67 with movement away from the perichromosomal layer to cytoplasmic dots that colocalize with nucleolin. These data indicate that pKi67 localization is regulated by the action of cell cycle-specific kinase(s) and phosphatase(s). The data presented here provide a starting point for the analysis of pKi67 function and regulation.
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PMID:Biochemical characterization of pKi67 with the identification of a mitotic-specific form associated with hyperphosphorylation and altered DNA binding. 1050 11

Cells require optimum protein synthetic activity in order to support cell proliferation, maintain homeostatic and metabolic integrity, and repair damage. Since growth depends on protein synthesis through ribosome biogenesis, the control of biosynthesis of ribosomes is necessarily a key element for control of growth. Nucleolin is a major nucleolar protein of exponentially growing eukaryotic cells, which is directly involved in the regulation of ribosome biogenesis and maturation. The highly conserved nucleolin contains three major domains through which it controls the organization of nucleolar chromatin, packaging of pre-RNA, rDNA transcription, and ribosome assembly. Numerous reports have implicated the involvement of nucleolin either directly or indirectly in the regulation of cell proliferation and growth, cytokinesis, replication, embryogenesis, and nucleogenesis. Nucleolin, an RNA binding protein, is also an autoantigen, a transcriptional repressor, and a switch region targeting factor. In addition, nucleolin exhibits autodegradation, DNA and RNA helicase activities, and DNA-dependent ATPase activity. An interesting aspect of nucleolin action is that it is a target for regulation by proteolysis, methylation, ADP-ribosylation, and phosphorylation by CKII, cdc2, PKC-xi, cyclic AMP-dependent protein kinase, and ecto-protein kinase. For these and other reasons, nucleolin is fundamental to the survival and proliferation of cells. Considerable progress has been made in recent years with the identification of new nucleolin binding proteins that may mediate these many nucleolin-dependent functions. Nucleolin also functions as a cell surface receptor, where it acts as a shuttling protein between cytoplasm and nucleus, and thus can even provide a mechanism for extracellular regulation of nuclear events. Exploration of the regulation of this multifaceted protein in a remarkable number of diverse functions is challenging.
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PMID:Molecular dissection of nucleolin's role in growth and cell proliferation: new insights. 1054 74

A 20-kDa DNA-binding protein that binds the AT-rich sequences within the promoters of the brain-specific protein kinase C (PKC) gamma and neurogranin/RC3 genes has been characterized as chromosomal nonhistone high-mobility-group protein (HMG)-I. This protein is a substrate of PKC alpha, beta, gamma, and delta but is poorly phosphorylated by PKC epsilon and zeta. Two major (Ser44 and Ser64) and four minor phosphorylation sites have been identified. The extents of phosphorylation of Ser44 and Ser64 were 1:1, whereas those of the four minor sites all together were <30% of the major one. These PKC phosphorylation sites are distinct from those phosphorylated by cdc2 kinase, which phosphorylates Thr53 and Thr78. Phosphorylation of HMG-I by PKC resulted in a reduction of DNA-binding affinity by 28-fold as compared with 12-fold caused by the phosphorylation with cdc2 kinase. HMG-I could be additively phosphorylated by cdc2 kinase and PKC, and the resulting doubly phosphorylated protein exhibited a >100-fold reduction in binding affinity. The two cdc2 kinase phosphorylation sites of HMG-I are adjacent to the N terminus of two of the three predicted DNA-binding domains. In comparison, one of the major PKC phosphorylation sites, Ser64, is adjacent to the C terminus of the second DNA-binding domain, whereas Ser44 is located within the spanning region between the first and second DNA-binding domains. The current results suggest that phosphorylation of the mammalian HMG-I by PKC alone or in combination with cdc2 kinase provides an effective mechanism for the regulation of HMG-I function.
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PMID:Phosphorylation of HMG-I by protein kinase C attenuates its binding affinity to the promoter regions of protein kinase C gamma and neurogranin/RC3 genes. 1061 44

Methylselenocysteine (MSC), an organic selenium compound is an effective chemopreventive agent against mammary cell growth both in vivo and in vitro but its mechanism of action is still not understood. We have previously demonstrated that MSC is able to inhibit growth in a synchronized TM6 mouse mammary epithelial tumor cell line at 16 h time point followed by apoptosis at 48 h. The decrease in cdk2 kinase activity was coincident with prolonged arrest of cells in S-phase. The present set of experiments showed that cdk2 phosphorylation was reduced by 72% in the MSC-treated cells at 16 h time point. Expression for gadd34, 45 and 153 was elevated 2.5 to 7 fold following MSC treatment only after 16 h time point. In order to investigate a possible upstream target for MSC, we analyzed protein kinase C (PKC) in this model. Total PKC activity was reduced in TM6 cells by MSC (50 microM) within 30 min of treatment, both in cytosolic (55.4 and 77.6%) and membrane (35.2 and 34.1%) fractions for calcium-dependent and independent PKCs, respectively. PMA significantly elevated the PKC activity in membrane fraction (P < 0.01) and MSC inhibited this activation by more than 57%. The effect of MSC was selenium specific as selenomethionine and sulfurmethyl-L-cysteine (SMC) did not alter PKC activity either in cytosolic or membrane fraction. Immunoblot analysis showed that PKC-alpha was translocated to the membrane by PMA and MSC did not alter this translocation. PKC-delta was faintly detectable in membrane fractions of control and MSC-treated cells. MSC treatment slightly reduced levels of PKC-e (in cytosolic and membrane fractions) and PKC-zeta (cytosolic fractions). The data presented herein suggest that PKC is a potential upstream target for MSC that may trigger one or all of the downstream effects; i.e. the decrease of cdk2 kinase activity, decreased DNA synthesis, elevation of gadd gene expression and finally apoptosis.
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PMID:Effects of methylselenocysteine on PKC activity, cdk2 phosphorylation and gadd gene expression in synchronized mouse mammary epithelial tumor cells. 1065 18

7-Hydroxystaurosporine (UCN-01), which was originally identified as a protein kinase C selective inhibitor, is currently in clinical trials as an anti-cancer drug. We previously showed that UCN-01 induced preferential G1-phase accumulation in tumor cells and this effect was associated with the retinoblastoma (Rb) protein and its regulatory factors, such as cyclin-dependent kinase 2 (CDK2) and CDK inhibitors p21Cip1/WAF1 and p27Kip1. We demonstrate here that G1-phase accumulation was induced by UCN-01 in Rb-proficient cell lines (WiDr and HCT116 human colon carcinomas and WI-38 human lung fibroblast), and it was accompanied by dephosphorylation of Rb. In addition, UCN-01-induced G1-phase accumulation was also demonstrated in a Rb-defective cell line (Saos-2 human osteosarcoma), but not in a simian virus 40 (SV40)-transformed cell line (WI-38 VA13). Apoptosis was induced by UCN-01 in the two Rb-deficient cell lines, but not in the other Rb-proficient cell lines. These observations suggest that G1-checkpoint function might be important for cell survival during UCN-01 treatment. In addition, there may be a UCN-01-responsive factor in the G1-checkpoint machinery other than Rb which is targeted by SV40. Further studies revealed a correlation between UCN-01-induced G1-phase accumulation and reduction of cellular CDK2 kinase activity. This reduction was strictly dependent on down-regulation of the Thr160-phosphorylated form of CDK2 protein, and coincided in part with up-regulation of p27Kip1, but it was independent of the level of the p21Cip1/WAF1 protein. These results suggest that G1-checkpoint function, including a CDK2-regulatory pathway, may be a significant determinant of the sensitivity of tumor cells to UCN-01.
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PMID:G1-checkpoint function including a cyclin-dependent kinase 2 regulatory pathway as potential determinant of 7-hydroxystaurosporine (UCN-01)-induced apoptosis and G1-phase accumulation. 1066 55


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