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)

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

Unfertilized frog eggs arrest at the second meiotic metaphase, due to cytostatic activity of the c-mos proto-oncogene (CSF). MAP kinase has been proposed to mediate CSF activity in suppressing cyclin degradation. Using an in vitro assay to generate CSF activity, and recombinant CL 100 phosphatase to inactivate MAP kinase, we confirm that the c-mos proto-oncogene blocks cyclin degradation through MAP kinase activation. We further show that for MAP kinase to suppress cyclin degradation, it must be activated before cyclin B-cdc2 kinase has effectively promoted cyclin degradation. Thus MAP kinase does not inactivate, but rather prevents the cyclin degradation pathway from being turned on. Using a constitutively active mutant of Ca2+/calmodulin dependent protein kinase II, which mediates the effects of Ca2+ at fertilization, we further show that the kinase can activate cyclin degradation in the presence of both MPF and the c-mos proto-oncogene without inactivating MAP kinase.
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PMID:MAP kinase does not inactivate, but rather prevents the cyclin degradation pathway from being turned on in Xenopus egg extracts. 883 8

Normal human cells from epidermis and from buccal mucosa were cultured to confluence in three media with graded differentiation potential (at low Ca2+, high Ca2+, and supplemented with serum) and treated with transforming growth factor beta 1 (TGF-beta 1), as had been done previously with interferon-gamma (IFN-gamma). The response of the cells to TGF-beta 1 was monitored in terms of the expression of regulatory genes associated with proliferation and differentiation (cdc2, c-myc, p53) and of genes for structural proteins expressed at varying stages of maturation (keratins K5 and K10, involucrin, flaggrin). For both tissues, the results obtained with both agents were very similar for those genes expressed in the basal cells (cdc2, c-myc, p53, K5), regardless of their function, but diverged for the other genes, which are expressed in the suprabasal cells. Another related contrast is that, although IFN-gamma induced apoptosis in epidermal keratinocytes cultured in the serum containing medium, TGF-beta 1 did not. Thus, the two agents appear to affect the earlier stages of cell differentiation in the same way but to differ at the later stages, particularly in that IFN-gamma pushes maturation further than does TGF-beta 1).
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PMID:Response of cultured cells from the epidermis and the buccal mucosa to TGF-beta 1 and comparison to interferon-gamma. 883 86

Glucose-regulated stress response of cancer cells occurs during the growth of solid tumors and is induced in culture by treatments with various agents, including 2-deoxyglucose, glucosamine, and calcium ionophore A23187. We previously reported that the three stressors commonly induced cell-cycle arrest in the G1 phase and resistance to antitumor drugs in human cancer A2780 and HT-29 cells. In this study, we investigated the mechanisms of stress-induced G1 arrest by determining the expression of cell-cycle-regulating proteins. Among G1 cyclins and cyclin-dependent kinases (cdk) examined, the expression levels of cyclin D1 preferentially decreased in the stressed cells. A time-course study showed that the decrease in cyclin D1 coincided with the appearance of hypophosphorylated retinoblastoma protein (pRb), which is the growth suppressive form. These findings suggest that the stress-induced G1 arrest is mediated through the down-regulation of cyclin D1-associated kinases (cdk4/6), pRb kinases during G1 phase. This was also supported by decreased cdk4 expression in stressed HT-29 cells. In addition, p21WAF1, a cdk inhibitor, was induced in the stressed cells, particularly A23187-treated cells. A23187, compared with the other stressors, caused extreme pRb hypophosphorylation, suggesting that p21WAf1 is involved in the regulation of pRb phosphorylation in the stressed cells. Our present findings could explain a molecular-based mechanism of a growth-arrested quiescent state and also resistance to chemotherapy of solid tumor cells.
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PMID:Glucose-regulated stresses cause decreased expression of cyclin D1 and hypophosphorylation of retinoblastoma protein in human cancer cells. 900 Jan 44

The effect of a calpain-selective cell permeant inhibitor, benzyloxycarbonyl Leu-Leu-Tyr diazomethylketone (ZLLY-CHN2), on the serum-stimulated growth of WI-38 human fibroblasts has been investigated. Only cell permeant protease inhibitors with activity against calpains prevented progression into S-phase. Protein blotting experiments indicated that p53 immunoreactivity increased in late G1 cells treated with ZLLY-CHN2. The content of p21Waf1/Cip1 CDK inhibitor also increased, providing a mechanism for the observed failure to enter S-phase. Further studies indicated that p53 could be degraded by a ZLLY-CHN2-sensitive protease immediately prior to S-phase, but that proteolysis did not occur after this critical time point. Chelation of extracellular Ca2+ by addition of EGTA inhibited the p53 degradation. Consistent with proteolysis of p53 in late G1 phase, mu-calpain immunoreactivity transiently accumulated in cell nuclei at this time. ZLLY-CHN2 did not appear to increase p53 mRNA in WI-38 cells. Purified mu-calpain required only 1 to 3 microM Ca2+ to proteolyze p53 in WI-38 cell lysates. These results indicate that ZLLY-CHN2 inhibits progression of WI-38 cells into S-phase by inactivating a calpain-like protease that is responsible for proteolysis of constitutively expressed p53 in late G1.
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PMID:Inhibition of the growth of WI-38 fibroblasts by benzyloxycarbonyl-Leu-Leu-Tyr diazomethyl ketone: evidence that cleavage of p53 by a calpain-like protease is necessary for G1 to S-phase transition. 901 11

In response to the neurohormone serotonin (5-hydroxytryptamine, 5-HT), prophase-arrested oocytes of the marine bivalve Crassostrea gigas (oyster) reinitiate meiosis, undergo germinal vesicle breakdown (GVBD), and are arrested again at metaphase I. We examined the pharmacological characteristics of 5-HT receptors and the signal transduction pathway following 5-HT stimulation in oyster oocytes. Among 5-HT agonists tested, only alpha-methyl 5-HT, a 5-HT2 agonist, induced GVBD although it was 1000 times less sensitive than 5-HT. The rank order of the potency of 5-HT antagonists to inhibit GVBD was propranolol, cyproheptadine > metoclopramide > mianserin. These results are quite different from those reported for other mollusks, suggesting the presence of unique 5-HT receptors on oyster oocytes. Using the fluorescent Ca2+ dyes fura 2 and calcium green and the pH indicator 1-hydroxypyrene-3,6,8-trisulfonic acid, we examined changes in intracellular Ca2+ ([Ca2+]i) and intracellular pH (pHi) during 5-HT-induced meiosis reinitiation. 5-HT did not trigger any changes in [Ca2+]i. However, an increase in pHi was observed during the 5-HT-induced meiosis reinitiation. The increased pHi level was rather small before GVBD and not necessary for GVBD, because lowering pHi by sodium acetate seawater (pH 7.0) did not prevent 5-HT-induced GVBD. Measurement of the kinase activity toward a peptide substrate specific to cdc2 demonstrated that maturation-promoting factor (MPF) was activated in accordance with the occurrence of GVBD in response to 5-HT. Therefore, it is likely that in oyster oocytes the signal transduction pathways and intracellular effectors participating in 5-HT-induced meiosis reinitiation via the activation of MPF are insensitive to [Ca2+]i and pHi.
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PMID:Change in intracellular Ca2+ is not involved in serotonin-induced meiosis reinitiation from the first prophase in oocytes of the marine bivalve Crassostrea gigas. 907 41

UCN-01 (7-hydroxyl-staurosporine) was originally isolated as a Ca2+- and phospholipid-dependent protein kinase C selective inhibitor and now is being developed as an anticancer agent. Results from our and other laboratories have suggested that UCN-01 induces preferential G1-phase accumulation in several human tumor cell lines tested. To elucidate this mechanism, we examined the effects of UCN-01 on several cell cycle-regulatory proteins critical for G1-S-phase transition in p53-mutated human epidermoid carcinoma A431 cells. After 24 h exposure at around 50% growth-inhibitory concentrations (IC50s), 260 and 520 nM, UCN-01 induced the accumulation of pRb (the dephosphorylated retinoblastoma protein form). The protein expression of cyclin A but not cyclin E was markedly reduced and that of cyclin D1 was partially reduced under the same condition. UCN-01 also showed the concentration-dependent inhibitions of the activity of cyclin-dependent kinase 2 (CDK2) using histone H1 and pRb as substrates in vitro (IC50, 530 and 640 nM, respectively). In addition, CDK2 activities of the cells pretreated with UCN-01 for 24 h at 260 and 520 nM were markedly inhibited, giving IC50s of far less than 260 nM. When the same cell lysates were analyzed by Western blotting for CDK2, the lower band (e.g., active and phosphorylated CDK2) was remarkably reduced, in accordance with the reduced activity. Furthermore, UCN-01 induced the expression of the CDK inhibitor p21 protein and its complex formation with CDK2 after 24 h exposure at 260 and 520 nM, whereas the expression level was very low or undetectable in untreated or DNA-damaged cells. The increase of p21 mRNA levels was also induced under the same condition. UCN-01 further increased luciferase activities in A431 cells transiently transfected with p21 promoter-luciferase reporter plasmid after 24 h exposure at 260 and 520 nM. UCN-01 also increased the expression of the CDK inhibitor p27 protein after 24 h exposure at 260 and 520 nM. These results suggest that G1-phase accumulation induced by UCN-01 is associated with dephosphorylation of Rb and CDK2 proteins as well as induction of CDK inhibitors p21 and p27.
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PMID:G1 phase accumulation induced by UCN-01 is associated with dephosphorylation of Rb and CDK2 proteins as well as induction of CDK inhibitor p21/Cip1/WAF1/Sdi1 in p53-mutated human epidermoid carcinoma A431 cells. 910 51

The involvement of cell cycle-regulatory proteins in apoptosis of neuronally differentiated PC12 cells induced by the removal of nerve growth factor and serum was examined. Three major findings are presented. (1) Cdc2 kinase protein levels increased fivefold in apoptotic PC12 cells by day 3 of serum and nerve growth factor deprivation. Histone H1 kinase activity was increased significantly in p13(suc1) precipitates of apoptotic PC12 cells, which was due to increased activation and/or expression of cdc2 kinase. (2) The protein levels of cyclin-dependent kinase 4, cyclin D, and proliferating cell nuclear antigen that are normally expressed in the cell cycle were increased during neuronal PC12 cell apoptosis. (3) The levels of the catalytic subunit, but not the regulatory subunit of the calcium/calmodulin-dependent protein phosphatase 2B, decreased significantly concomitant with a significant decrease in protein phosphatase 2B activity early in the apoptotic process. Protein phosphatase 2A activity decreased slightly but significantly after 3 days of serum and nerve growth factor deprivation, and no alterations in protein phosphatase 1 were observed during the apoptotic process. These data demonstrate that certain cell cycle-regulatory proteins are inappropriately expressed and that alterations in specific phosphorylation events, as indicated by the increase in histone H1 kinase activity and the decrease in protein phosphatase 2B activity, are most likely occurring during apoptosis of PC12 cells. These observations support the hypothesis that apoptosis may be due in part to a nondividing cell's uncoordinated attempt to reenter and progress through the cell cycle.
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PMID:Select alterations in protein kinases and phosphatases during apoptosis of differentiated PC12 cells. 916 26

Combinations of epidermal growth factor (EGF) and KCl stimulate differentiation in PC12 cells, independent of extracellular calcium [Mark et al., Stimulation of neurite outgrowth in PC12 cells by EGF and KCl depolarization: a Ca2+-independent phenomenon, J. Cell Biol., 130 (1995) 701-710]. Since EGF is a proliferative agent that normally does not stimulate differentiation of PC12 cells, we hypothesize that KCl plus EGF may cause differentiation because of the anti-proliferative activity of KCl. Here we report that treatment of PC12 cells with KCl plus EGF resulted in a significant decrease in proliferation and DNA synthesis compared with cells treated with EGF alone. In addition, KCl significantly reduced the EGF-induced expression of cell cycle progression factors cdk2, cdk4, cyclin B1 and PCNA. These data suggest that the anti-proliferative activity of KCl may convert EGF from a proliferative factor to a progression factor.
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PMID:Antiproliferative activity of KCl contributes to EGF-induced neurite outgrowth in PC12 cells. 925 67

Disassembly of the sperm nuclear envelope at fertilization is one of the earliest events in the development of the male pronucleus. We report that nuclear lamina disassembly in interphase sea urchin egg cytosol is a result of lamin B phosphorylation mediated by protein kinase C (PKC). Lamin B of permeabilized sea urchin sperm nuclei incubated in fertilized egg G1 phase cytosolic extract is phosphorylated within 1 min of incubation and solubilized prior to sperm chromatin decondensation. Phosphorylation is Ca2+-dependent. It is reversibly inhibited by the PKC-specific inhibitor chelerythrine, a PKC pseudosubstrate inhibitor peptide, and a PKC substrate peptide, but not by inhibitors of PKA, p34(cdc2) or calmodulin kinase II. Phosphorylation is inhibited by immunodepletion of cytosolic PKC and restored by addition of purified rat brain PKC. Sperm lamin B is a substrate for rat brain PKC in vitro, resulting in lamin B solubilization. Two-dimensional phosphopeptide maps of lamin B phosphorylated by the cytosolic kinase and by purified rat PKC are virtually identical. These data suggest that PKC is the major kinase required for interphase disassembly of the sperm lamina.
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PMID:Protein kinase C-mediated interphase lamin B phosphorylation and solubilization. 926 Nov 38


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