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Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In both starfish and amphibian oocytes, the activity of a major protein kinase which is independent of Ca2+ and cyclic nucleotides increases dramatically at meiotic and mitotic nuclear divisions. The in vivo substrates of this kinase are unknown, but phosphorylation of H1 histone can be used as an in vitro assay. We have purified this kinase from starfish oocytes. The major band in the most highly purified preparation contained a polypeptide of relative molecular mass (Mr) 34,000 (34K). This is the same size as the protein kinase encoded by cdc2+, which regulates entry into mitosis in fission yeast and is a component of MPF purified from Xenopus. Here, we show that antibodies against p34 recognize the starfish 34K protein and propose that entry into meiotic and mitotic nuclear divisions involves activation of the protein kinase encoded by a homologue of cdc2+. Given the wide occurrence of cdc2+ homologues from budding yeast to Xenopus and human cells, this activation may act as a common mechanism controlling entry into mitosis in eukaryotic cells.
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PMID:Activation at M-phase of a protein kinase encoded by a starfish homologue of the cell cycle control gene cdc2+. 341 86

Changes in the extent of protein phosphorylation and their possible correlation with changes in the activity of maturation-promoting (MPF) factor were investigated throughout meiotic maturation and following activation of amphibian and starfish oocytes. Despite several exceptions in the pattern of phosphorylation of individual proteins, high and low levels of protein phosphorylation were found to be correlated with high and low levels of MPF activity. Both the extent of protein phosphorylation and MPF activity were found to drop upon parthenogenetic activation and to cycle synchronously thereafter in the amphibian. In contrast no drop in MPF activity or in the extent of protein phosphorylation was observed following activation of starfish oocytes with ionophore A23187. This suggests that changes of protein phosphorylation and of MPF activity are rather related to the progression of the cell cycle than directly to Ca2+-dependent activation reaction. In amphibians global protein kinase activity in homogenates was found to drop with MPF activity following activation. Changes in the ratio of threonine vs serine phosphorylation were also investigated during the course of meiotic maturation and activation in both amphibian and starfish oocytes: changes in the activity of MPF were found to be better correlated with changes in threonine than serine phosphorylation.
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PMID:Changes in the activity of the maturation-promoting factor during meiotic maturation and following activation of amphibian and starfish oocytes: their correlations with protein phosphorylation. 352 14

The availability of the pure inhibitor of cAMP-dependent protein kinase prompted a re-examination of the inhibitor-induced meiotic maturation of Xenopus laevis oocytes. Injection of the inhibitor (1.5 microM) triggered 100% germinal vesicle breakdown faster than progesterone and slower than the maturation-promoting factor: at 0.15 microM, the inhibitor still triggered 100% meiosis, but with a much slower kinetics. In contrast, injection of 24 microM calmodulin resulted in less than 50% GVBD, and results were variable from female to female. Combined injection of inhibitor and calmodulin failed to show any synergism, which does not favour hypotheses according to which calmodulin acts by activation of cyclic nucleotide phosphodiesterase. The net effect of the inhibitor is to decrease the concentration of the free catalytic sub-unit of cAMP-dependent protein kinase, fully dissociated in the unstimulated oocyte, as shown by the absence of effect of pretreatment with cholera toxin on the inhibitor-induced maturation. After such decrease by about 1 microM, a maturation protein, Mp-P, is dephosphorylated by phosphoprotein phosphatases. Dephospho-Mp triggers the synthesis of MPF in cycloheximide-sensitive steps. Finally, MPF triggers GVBD in steps insensitive to cycloheximide. Evidence for such a 4-step scheme--fall in cAMP levels, then in C sub-unit levels, dephosphorylation of Mp leading to the synthesis of MPF and finally MPF-triggered GVBD--is presented and discussed.
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PMID:The pure inhibitor of cAMP-dependent protein kinase initiates Xenopus laevis meiotic maturation. A 4-step scheme for meiotic maturation. 701 32

Brefeldin A, a fungal metabolite which disrupts protein traffic, provokes indirect activation of cdc2 protein kinase in Xenopus oocytes. Cdc2 protein kinase activation was judged by MPF (M-phase factor) transfer activity, histone H1 kinase activity, and phosphorylation in vivo of the guanine-nucleotide exchange complex EF-1 beta gamma delta. Oocytes resumed complete meiosis upon brefeldin A treatment. Cdc2 protein kinase, MAP kinase, cyclin B, MPF, and protein synthesis changes were all comparable in brefeldin A-treated oocytes and in progesterone-induced oocytes. ED50 for brefeldin A was 0.6 microM. Brefeldin A activation of cdc2 protein kinase occurs with a long time course. Simultaneous treatment of the oocytes at a subthreshold concentration of 1 nM progesterone and 30 microM brefeldin A considerably shortened the kinetics of maturation. Brefeldin A induction of maturation was sensitive to drugs that act on cAMP metabolism. ID50 for IBMX was 0.1 mM, compared to 1 mM for progesterone-treated oocytes. Brefeldin A inhibited protein traffic in oocytes as determined from protein export experiments. ID50 was between 0.1 and 1 microM. Our results give new insights into the possible mechanism of induction of meiotic maturation and further demonstrate that brefeldin A acts on cell cycle regulatory elements.
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PMID:Brefeldin A provokes indirect activation of cdc2 kinase (MPF) in Xenopus oocytes, resulting in meiotic cell division. 754 76

Microinjection of bacterially expressed human cdc25A protein into Xenopus prophase oocytes provokes the activation of p34cdc2 kinase and the tyrosine dephosphorylation of p34cdc2 in the presence or absence of protein synthesis. The level of p34cdc2 kinase activity then drops in parallel with the degradation of cyclin B2 and finally increases again to stabilize at a high level. Cdc25 microinjection induces the assembly of a metaphase I spindle which is abnormally located in the deep cytoplasm. Moreover, oocytes arrest at the metaphase I stage and do not reach metaphase II even 10 h after cdc25 microinjection. The extended metaphase I period observed in cdc25-injected oocytes results from an equilibrium between degradation of cyclins and synthesis of new cyclins. This is in contrast with progesterone-stimulated oocytes where cyclin degradation is turned off when oocytes enter metaphase II. During metaphase I, the reactivation of MPF activity can be disrupted in two different ways: 1) cycloheximide, an inhibitor of protein synthesis, by preventing the synthesis of new cyclins, provokes the disappearance of MPF kinase activity and the reformation of a nucleus; 2) when the cAMP level is increased during the metaphase I period in cdc25-injected oocytes, MPF kinase activity drops following a rephosphorylation of tyrosine 15 of p34cdc2, while the cyclin turn-over remains unaffected. Moreover, increasing the cAMP level in prophase oocytes totally prevents the action of cdc25. Our results indicate that in Xenopus oocytes, the PKA pathway negatively regulates the activation of MPF and the activity of p34cdc2/cyclin B complex through tyrosine phosphorylation of p34cdc2 during metaphase I.
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PMID:Microinjection of Cdc25 protein phosphatase into Xenopus prophase oocyte activates MPF and arrests meiosis at metaphase I. 773 15

A universal model of the control of the cell cycle in eukaryotic organisms has emerged from the discovery that MPF (maturation or mitosis promoting factor) is a heterodimer consisting of a catalytic subunit (p34cdc2) and a regulatory subunit (mitotic cyclin) encoded by a pair of conserved genes. A prominent feature of the periodic activation of the protein kinase p34cdc2 is the gradual accumulation of cyclin in interphase and its abrupt degradation in mitosis, which is believed to be required for inactivation of MPF and exit from mitosis. Utilizing the precise natural synchrony of mitosis of the plasmodium of the myxomycete Physarum, the high affinity of the p34cdc2/cyclin B complex to p13suc1 Sepharose beads, and immunological reagents including three different anticyclin B antibodies and the anti-PSTAIR antibody, a transient histone H1 kinase activation but not fluctuation in the abundance of cyclin B have been detected during mitosis. It is argued that cyclin degradation may be required for cytokinesis and/or postmitotic controls of cell proliferation in G1 phase and cell-to-cell signaling in development but not for the inactivation of histone H1 kinase in mitosis.
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PMID:A non-cycling mitotic cyclin in the naturally synchronous cell cycle of Physarum polycephalum. 788 99

HeLa cells in G2 phase are temporarily inhibited and prevented from entering mitosis by treatment with the phorbol ester TPA (12-O-tetradecanoylphorbol-13-acetate), whereas cells in mitosis are refractory to TPA and divide. In this study the possibility was tested that TPA may interfere with the regulatory cycle of MPF (mitosis promoting factor), the rate-limiting protein kinase for cell division. MPF, consisting of the catalytic subunit p34cdc2 and the regulatory subunit Cyclin B, is known to be activated at the transition from G2 phase to mitosis through dephosphorylation at Tyr15 and to become inactivated after metaphase by proteolysis. Treatment of HeLa cells (synchronized around the G2-M transition) with TPA (10(-7) M) has now been shown to induce an overall decrease of the histone H1 kinase activity associated with anti-p34cdc2 immunoprecipitates after about 20 to 30 min. In metaphase cells, the histone H1 kinase activity of p34cdc2 was shown to remain unaffected by TPA treatment. In cultures enriched in G2 cells neither the amount of p34cdc2 protein nor that of Cyclin B was influenced by TPA. Moreover, the p34cdc2/Cyclin B complex formation was also unaffected. However, p34cdc2 from cultures treated with TPA was more intensely stained by anti-phosphotyrosine antibodies than that of control cells, indicating that TPA treatment probably prevented the tyrosine dephosphorylation required for expression of the histone H1 kinase activity of the complex. The results indicate that TPA treatment of HeLa cultures rapidly stops the G2-M transition because it very rapidly prevents the p34cdc2/Cyclin B complex in G2 cells from developing histone H1 kinase activity.
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PMID:Phorbol ester TPA rapidly prevents activation of p34cdc2 histone H1 kinase and concomitantly the transition from G2 phase to mitosis in synchronized HeLa cells. 818 33

During studies of the activation and inactivation of the cyclin B-p34cdc2 protein kinase (MPF) in cell-free extracts of Xenopus oocytes and eggs, we found that a bacterially expressed fusion protein between the Escherichia coli maltose-binding protein and the Xenopus c-mos protein kinase (malE-mos) activated a 42 kDa MAP kinase. The activation of MAP kinase on addition of malE-mos was consistent, whereas the activation of MPF was variable and failed to occur in some oocyte extracts in which cyclin A or okadaic acid activated both MPF and MAP kinase. In cases when MPF activation was transient, MAP kinase activity declined after MPF activity was lost, and MAP kinase, but not MPF, could be maintained at a high level by the presence of malE-mos. When intact oocytes were treated with progesterone, however, the activation of MPF and MAP kinase occurred simultaneously, in contrast to the behaviour of extracts. These observations suggest that one role of c-mos may be to maintain high MAP kinase activity in meiosis. They also imply that the activation of MPF and MAP kinase in vivo are synchronous events that normally rely on an agent that has still to be identified.
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PMID:The c-mos proto-oncogene protein kinase turns on and maintains the activity of MAP kinase, but not MPF, in cell-free extracts of Xenopus oocytes and eggs. 838 16

Cell cycle progression in cycling Xenopus egg extracts is accompanied by fluctuations in the concentration of adenosine 3',5'-monophosphate (cAMP) and in the activity of the cAMP-dependent protein kinase (PKA). The concentration of cAMP and the activity of PKA decrease at the onset of mitosis and increase at the transition between mitosis and interphase. Blocking the activation of PKA at metaphase prevented the transition into interphase; the activity of M phase-promoting factor (MPF; the cyclin B-p34cdc2 complex) remained high, and mitotic cyclins were not degraded. The arrest in mitosis was reversed by the reactivation of PKA. The inhibition of protein synthesis prevented the accumulation of cyclin and the oscillations of MPF, PKA, and cAMP. Addition of recombinant nondegradable cyclin B activated p34cdc2 and PKA and induced the degradation of full-length cyclin B. Addition of cyclin A activated p34cdc2 but not PKA, nor did it induce the degradation of full-length cyclin B. These findings suggest that cyclin degradation and exit from mitosis require MPF-dependent activation of the cAMP-PKA pathway.
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PMID:Requirement for cAMP-PKA pathway activation by M phase-promoting factor in the transition from mitosis to interphase. 859 31

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


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