Isolation of histone kinases.
Explore the source record for details and available documents.
Cell Line↗
SEARCH · Search PubMed
Results for “Protamine Kinase”
Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
Differential effects of polyamines on rat thyroid protein kinase activities.
Ornithine decarboxylase, the rate-limiting enzyme in polyamine biosynthesis, has been shown to be regulated in thyroid by thyrotropin both in vivo and in vitro. Little, however, is known of the role of polyamines in thyroid cell function. Since studies in other tissues suggest that polyamines may influence protein phosphorylation, we studied the effect of the polyamines on various protein kinase activities in rat thyroid. Putrescine, spermidine, and spermine inhibit cyclic-AMP-dependent histone H1 kinase activity when measured in the cytosol fraction of rat thyroid; this effect is largely reproduced by NaCl concentrations of equivalent ionic strength. Both spermidine and spermine effect a 1.6-2.4-fold increase in cytosolic cyclic-AMP-independent (messenger-independent) casein kinase activity; stimulation by both polyamines is maximal at 5mM. A similar profile of stimulation is observed for messenger-independent casein kinase activity in crude nuclear preparations. Sodium chloride fails to stimulate both cytosolic and nuclear messenger-independent casein kinase activities at ionic strength equivalent to the spermine concentrations used. Spermine, but not putrescine, spermidine, or sodium chloride, inhibits calcium/phospholipid-dependent protein kinase C activity in cytosol extracts partially purified by DEAE chromatography. These findings suggest that regulation of protein kinase(s) by polyamines may represent a proximal locus (i) of action of thyrotropin-regulated ornithine decarboxylase activity in thyroid.
Animals↗
Differential regulation of the synthesis and activity of the major cyclin-dependent kinases, p34cdc2, p33cdk2, and p34cdk4, during cell cycle entry and progression in normal human T lymphocytes.
Three major cyclin-dependent kinases, p34cdc2, p33cdk2, and p34cdk4 were examined in normal human T cells stimulated to enter the cell cycle in vitro. None of the three genes was expressed in resting T cells. Transcripts form the cdk4 and cdk2 genes were detectable as early as 3 and 8 hr after stimulation, respectively, whereas cdc2 gene transcripts were not detectable until about 24 hr, shortly before S phase entry. Immunoblot analysis showed that resting T cells contained little p34cdk4, no p34cdc2, and a low level of p33cdk2 protein. Increased amounts of p34cdk4, p33cdk2, and p34cdc2 proteins were seen at about 7, 10, and 30 hr after stimulation, respectively. Immunoprecipitates of each of the kinases were assessed for histone H1 kinase activity. Activity due to p33cdk2 first became detectable in mid-G1 phase and increased dramatically after entry into S phase. Active p34cdc2 kinase was not detected until about 40 hr after stimulation, about 10 hr after the first appearance of the protein. Immunoprecipitates of p34cdk4 possessed almost no H1 histone kinase activity; however, activity was detected as early as 10 hr after cell activation when a protein (p60Rb) derived from the retinoblastoma susceptibility gene product was used as substrate. Cells were synchronized about the G1/S and G2/M borders by aphidicolin and nocodazole. Cells arrested prior to S-phase contained high levels of active p33cdk2 and essentially no active p34cdc2, despite the fact that large amounts of both proteins were present. Cells arrested by nocodazole had high levels of active p34cdc2 and greatly reduced levels of p33cdk2 kinase activity. The results suggest that the major role for the p34cdc2 kinase is at mitosis, whereas that for p33cdk2 is in late G1 and/or S phase. The p34cdk4 protein, present in aphidicolin-blocked cells, was nearly absent from cells arrested at the G2/M border; however, kinase activity was low in cells blocked at both points, suggesting that the major role for p34cdk4 may be in G1 phase.
CDC2 Protein Kinase↗
Cyclin A and Cdk2 kinase activity are suppressed by combined treatment with tumor necrosis factor-alpha and interferon-alpha.
We previously reported that combined treatment with tumor necrosis factor-alpha (TNF-alpha) and interferon-alpha (IFN-alpha) showed a synergistic antitumor effect via regulation of cell cycle progression in the S phase. Here, we investigated the effect of the combined treatment with TNF-alpha and IFN-alpha on cell cycle regulating protein in RPMI 4788 cells. Treatment with TNF-alpha or IFN-alpha alone showed no effect on these proteins, however, the combined treatment showed suppression of cyclin A protein and its associated kinase activity. Furthermore, although the combined treatment inhibited Cdk2 kinase activity, the amount of Cdk2 protein was not affected. These results suggested that TNF-alpha and IFN-alpha work together to suppress cyclin A and Cdk2 kinase activity and to inhibit cell cycle progression in the S phase.
CDC2 Protein Kinase↗
Phorbol ester TPA rapidly prevents activation of p34cdc2 histone H1 kinase and concomitantly the transition from G2 phase to mitosis in synchronized HeLa cells.
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.
CDC2 Protein Kinase↗
Expression of the catalytic domain of cyclic GMP-dependent protein kinase in a baculovirus system.
The Type I cGMP-dependent protein kinase catalytic domain (residues 336-671 from the I alpha isoform) has been expressed as a cGMP independent kinase in a baculovirus system. Using peptide substrates, the protein retains similar substrate specificity as the native holoenzyme. The recombinant catalytic domain catalyzes the phosphorylation of histone, but does not display the inhibition using non-substrate histones which has been described for the holoenzyme. The catalytic domain is an active kinase in mammalian cells also since vascular smooth muscle cells transfected with the cDNA encoding the catalytic domain display altered morphology. The catalytic domain of G-kinase may be a useful tool for delineating the role of cGMP-mediated protein phosphorylation in cell systems.
Amino Acid Sequence↗
Histone phosphorylation in phorbol ester stimulated and beta-adrenergically stimulated mouse epidermis in vivo and characterization of an epidermal protein phosphorylation system.
Under certain physiological conditions a change in the phosphorylation of histones in mouse epidermis in vivo was observed. Thus a single local application of the tumor-promoting mitogen 12-O-tetradecanoylphorbol-13-acetate caused a long-lasting increase of histone H1 phosphorylation which paralleled stimulated cell proliferation. Injection of the antimitotic beta-adrenergic agonist isoproterenol led to a temporary decrease in the rate of phosphorylation of H1, H2A and H2B immediately after cyclic AMP accumulation. A complete protein phosphorylation system could be demonstrated in mouse epidermis homogenates. The following enzyme activities were partially purified and characterized: a cyclic AMP-dependent histone kinase; a 'casein kinase' and an 'unspecific' protein kinase; a histone-specific protein phosphatase; and two 'unspecific' phosphoprotein phosphatases. In addition, a stimulatory effect of cyclic GMP on histone phosphorylation was observed. The enzymes were found to be predominantly localized in the 105000 X g supernatant, but a small proportion of protein kinase and phosphatase activity could be regularly demonstrated in cell nuclei.
Animals↗
Preparing the target for the bullet.
Explore the source record for details and available documents.
Animals↗
The kinetics of H1 histone kinase activation during the cell cycle of wild-type and wee mutants of the fission yeast Schizosaccharomyces pombe.
H1 histone kinase activity has been followed in selection-synchronised cultures of fission yeast wild-type and wee1 mutant cells, and in induction-synchronised cells of the mutant cdc2-33. The main conclusions are: (1) in all three cases, the peak of activity is near mitosis. (2) The rise in activity is relatively slow starting in wild type at 0.4 of the cycle before mitosis. It is proposed that the beginning of the rise is the first identified event in the mitotic control. (3) The rise is twice as fast in wee and starts nearer to mitosis. (4) In all cases the beginning of the rise is in G2. (5) The fall in activity is also slow, lasting for 0.25 of the cycle, in wild type. Exit from mitosis happens well before activity has fallen to baseline. (6) In a range of size mutants, activity is roughly proportional to cell size. It is suggested that the kinase may have a cytoplasmic function. (7) Estimates have been made of the timing of mitosis in the mutants. In wee, mitosis is 0.14 of the cycle earlier than in wild type because the cells have a longer septated period at the end of the cycle. (8) A novel method has been developed for eliminating the effects of the partial asynchrony in synchronous cultures, without which the kinetic analysis would have been inaccurate.
CDC2 Protein Kinase↗
[Protein kinase activity in proliferating Ehrlich ascites tumor cells].
It was demonstrated that the activity of cAMP-dependent histone and casein kinases is maximal at the end of the exponential phase of tumour growth, i. e., on the 8-10th day and decreases in the stationary growth phase, i. e., by the 15th day. The maximal activity of histone and casein kinases is observed in the S-phase of the cell cycle, the degree of cAMP-dependent histone kinase activation reaching its maximum. A chromatographic analysis of cytosolic protein kinases revealed a significant increase in the activity of cAMP-dependent histone kinase of the second type within the same time interval. Cell proliferation was paralleled with changes in the relative content of the two forms of casein kinases.
Animals↗
Characterization of the in vitro reconstituted cyclin A or B1-dependent cdk2 and cdc2 kinase activities.
Human cyclins A and B1 were assembled with the cdk2 or cdc2 protein to reconstitute their respective kinase activities in vitro. Both cyclins complemented either cdk2 or cdc2, yielding kinase activities that supported the phosphorylation of histone H1. Activation of cdk2-catalyzed H1 kinase activity by cyclin A required a 10-min preincubation of the two components, whereas cdc2 kinase supported phosphate incorporation without a detectable time lag upon the addition of cyclin B1, suggesting a slower association rate of cdk2 with cyclin A compared with cdc2 and cyclin B1. Both cdk2 and cyclin A, as well as cdc2 and cyclin B1, formed stable complexes in the absence of ATP and substrate that could be isolated after glycerol gradient centrifugation. Incubation of the isolated complexes with ATP and histone H1 supported the phosphorylation of the substrate. Cyclin A-activated cdk2 or cdc2 phosphorylated p107, a pRB-related cellular protein, 10 times more effectively than the cyclin B1-complexed kinases. This was most likely due to a direct association of cyclin A with p107 (Ewen, M. E., Faha, B., Harlow, E., and Livingston, D. (1992) Science 255, 85-87; Faha, B., Ewen, M. E., Tsai, L.-H., Livingston, D., and Harlow, E. (1992) Science 255, 87-90). The reconstituted cdc2-cyclin B1 complex incorporated 4-5-fold more phosphate into the p34 subunit of the three-subunit (p70, p34, and p14) human single-stranded DNA-binding protein (also called RP-A), a DNA replication and DNA repair factor, than cdc2-cyclin A. No detectable phosphorylation of the p34 protein was observed with cdk2 complexed with either cyclin B1 or A. These data indicate that both cyclins as well as the catalytic subunits are important factors in controlling the rate of phosphorylation of a given substrate. The cyclin-activated cdc2 family kinases may target their cellular substrates through cyclin-mediated protein-protein interactions.
CDC2 Protein Kinase↗
Protein kinase activities during early development of Ascaris suum.
Explore the source record for details and available documents.
Animals↗
New evidence for compartmentalization of ovarian 3',5'-cyclic AMP-dependent protein kinase activity.
Explore the source record for details and available documents.
Animals↗
Reversible tyrosine phosphorylation of cdc2: dephosphorylation accompanies activation during entry into mitosis.
Tyrosine phosphorylation of cdc2 is regulated in the cell cycle of mouse 3T3 fibroblasts. Phosphotyrosine in cdc2 is detectable at the onset of DNA synthesis and becomes maximal in the G2 phase of the cell cycle. Quantitative tyrosine dephosphorylation of cdc2 occurs during entry into mitosis and no phosphotyrosine is detected during the G1 phase of the cell cycle. While increasing tyrosine phosphorylation of cdc2 correlates with the formation of a cdc2/p62 complex, the tyrosine phosphorylated cdc2 is inactive as a histone H1 kinase. cdc2 is fully dephosphorylated in its most active mitotic form, yet specific tyrosine dephosphorylation of interphase cdc2 in vitro is insufficient to activate the kinase. In vivo inhibition of tyrosine dephosphorylation by exposure of cells to a phosphatase inhibitor is associated with G2 arrest, which is reversible upon the removal of the phosphatase inhibitor. Tyrosine dephosphorylation of cdc2 may be one of a number of obligatory steps in the mitotic activation of the kinase.
Animals↗
Some characteristics and hormonal control of thyroidal cAMP-independent protein kinases.
Cyclic AMP-independent protein kinases in cytosol from rat thyroid glands were evaluated using histone and casein as exogenous substrates. Compared with other rat tissues, thyroid gland is rich in histone kinases, while its casein kinase activity is lower than that of liver and brain. Thyroidal cAMP-independent protein kinases can be resolved by sucrose gradient ultracentrifugation into two distinct peaks of histone (HKi-1 and HKi-2) and two peaks of casein (CKi-1 and CKi-2) kinases. An intermediate peak of histone kinase activity is only occasionally present. The four main kinase peaks are distinct with respect to several properties: their sedimentation coefficients are significantly different; only one out of the four peaks (CKi-2) can use GTP as substrate; monovalent ions strongly inhibit (50%) light peaks (HKi-1 and CKi-1), while heavy peaks (HKi-2 and CKi-2) are slightly but significantly stimulated (30%); light peaks are very sensitive to thermal inactivation, while heavy peaks are much more resistant. Their reactivity to hormonal action is different: in chronically stimulated glands HKi-2 is selectively and strongly stimulated (240%) while CKi-1 is not changed. In human pathological tissues independent alterations in different kinase entities were observed compared with healthy tissue. In conclusion, the four thyroidal c-AMP-independent protein kinases resolved on sucrose gradient seem to represent distinct entities which are independently and selectively controlled by hormones, and specifically altered in human pathological tissues.
Animals↗
Induction of a G2-phase arrest in Xenopus egg extracts by activation of p42 mitogen-activated protein kinase.
Previous work has established that activation of Mos, Mek, and p42 mitogen-activated protein (MAP) kinase can trigger release from G2-phase arrest in Xenopus oocytes and oocyte extracts and can cause Xenopus embryos and extracts to arrest in mitosis. Herein we have found that activation of the MAP kinase cascade can also bring about an interphase arrest in cycling extracts. Activation of the cascade early in the cycle was found to bring about the interphase arrest, which was characterized by an intact nuclear envelope, partially condensed chromatin, and interphase levels of H1 kinase activity, whereas activation of the cascade just before mitosis brought about the mitotic arrest, with a dissolved nuclear envelope, condensed chromatin, and high levels of H1 kinase activity. Early MAP kinase activation did not interfere significantly with DNA replication, cyclin synthesis, or association of cyclins with Cdc2, but it did prevent hyperphosphorylation of Cdc25 and Wee1 and activation of Cdc2/cyclin complexes. Thus, the extracts were arrested in a G2-like state, unable to activate Cdc2/cyclin complexes. The MAP kinase-induced G2 arrest appeared not to be related to the DNA replication checkpoint and not to be mediated through inhibition of Cdk2/cyclin E; evidently a novel mechanism underlies this arrest. Finally, we found that by delaying the inactivation of MAP kinase during release of a cytostatic factor-arrested extract from its arrest state, we could delay the subsequent entry into mitosis. This finding suggests that it is the persistence of activated MAP kinase after fertilization that allows the occurrence of a G2-phase during the first mitotic cell cycle.
Animals↗
Activation of M-phase-specific histone H1 kinase by modification of the phosphorylation of its p34cdc2 and cyclin components.
An M-phase-specific histone H1 kinase (H1K) has been described in a wide variety of eukaryotic cell types undergoing the G2/M transition in the cell division cycle. We have used p13suc1-Sepharose affinity chromatography to purify H1K to near homogeneity from matured starfish oocytes. A yield of 67% was obtained. Active H1K behaves as a 90- to 100-kD protein and appears to be constituted of equimolar amounts of cyclin and p34cdc2. The p34cdc2 subunit becomes tyrosine-dephosphorylated as the H1K is activated during entry of the oocytes into M phase, whereas the cyclin subunit is reciprocally phosphorylated. Acid phosphatase treatment of inactive p34cdc2/cyclin complex induces p34cdc2 dephosphorylation and three- to eightfold stimulation of the enzyme activity. These results suggest that active M-phase-specific H1K is constituted of both dephosphorylated p34cdc2 and phosphorylated cyclin.
Acid Phosphatase↗
Specific regulation by steroid hormones of protein kinases in the endometrium. 2. Alteration in levels of protein kinases in human endometrium during the menstrual cycle.
The alteration in the levels of multiple protein kinases has been studied in human endometrium during the menstrual cycle. Protein kinases were examined in the cytosol fraction and separated by DEAE-cellulose column chromatography. The level of type I adenosine 3',5'-monophosphate-dependent (cAMP-dependent) protein kinase was higher than that of type II cAMP-dependent protein kinase at the proliferative phase. At the late secretory phase, type I cAMP-dependent protein kinase markedly decreased, whereas type II cAMP-dependent protein kinase remained at the same level. Therefore, type II cAMP-dependent protein kinase was exclusively predominant at this phase. cAMP-dependent and cAMP-independent protein kinases with protamine increased more at the late secretory phase than at the proliferative phase. The results presented demonstrate a specific alteration in protein kinase activities during the menstrual cycle and are compatible with those obtained in rabbit endometrium (the preceding paper).
Adult↗