共查询到20条相似文献,搜索用时 15 毫秒
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Coupling of the group I metabotropic glutamate receptors, mGlu1a and mGlu5a, to the cAMP response element binding protein (CREB) has been studied in Chinese hamster ovary cell lines where receptor expression is under the control of an inducible promoter. Both receptors stimulate CREB phosphorylation with similar time courses, and agonist potency was also comparable between the two receptors. Stimulation of cells in Ca(2+)-free medium containing EGTA (100 microm), with or without the additional depletion of intracellular stores, caused marked decreases in agonist-mediated responses in both cell lines. Down-regulation of protein kinase C (PKC) activity by phorbol ester treatment, or treatment with the broad spectrum PKC inhibitor Ro 31-8220, partially attenuated both mGlu1a and mGlu5a receptor-mediated responses. Furthermore, stimulation of cells in the absence of extracellular Ca(2+) following prior PKC down-regulation resulted in additive inhibitory effects. The involvement of extracellular signal-regulated kinases (ERK1/2), Ca(2+)/calmodulin or Ca(2+)/calmodulin-dependent protein kinases was assessed using pharmacological inhibitors. Results indicated that coupling of the group I mGlu receptors to CREB phosphorylation occurs independently of these pathways. Thus, although the [Ca(2+)](i) signatures activated by these mGlu receptors differ, they couple to CREB with comparable potency and recruit similar downstream components to execute CREB phosphorylation. 相似文献
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Chang JH Vuppalanchi D van Niekerk E Trepel JB Schanen NC Twiss JL 《Journal of neurochemistry》2006,99(6):1517-1530
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Carlos J. Ciudad Jordi Vila M.Angels Mor Joan J. Guinovart 《Molecular and cellular biochemistry》1987,73(1):37-44
Extracts obtained from rat hepatocytes incubated with saline, glucagon or insulin were electrophoresed on polyacrylamide gels and then assayed for cyclic (3H)AMP binding capacity. Analysis of the binding patterns demonstrated that glucagon dissociated a holoenzyme of cyclic AMP-dependent protein kinase in a dose-dependent manner. The increase in free regulatory subunits and, hence, in free catalytic subunits explains the activation of this enzyme by glucagon in the liver. Insulin decreased both the amount of cyclic (3H)AMP bound to the holoenzyme and the capacity of the enzyme to be dissociated when the extracts were incubated with increasing concentrations of this cyclic nucleotide. We propose that these insulin-induced effects are determined by an inhibition of the cyclic AMP binding capacity of this protein kinase. This mechanism could account for the inactivation of cyclic AMP-dependent protein kinase that insulin causes in the liver.Abbreviations cAMP
(cyclic AMP), Adenosine 3,5 monophosphate
- (3H)cAMP
cyclic (3H)AMP
- MIX
1-methyl-3-isobutylxanthine 相似文献
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Extracellular signal-regulated kinases, protein kinase B/Akt and cyclase response element-binding protein play important roles in drug-induced neuroadaptations. Acute psychostimulant exposure rapidly alters the phosphorylation of these proteins in the striatum but less is known about their responses to repeated stimulant administration. In this study the phosphorylated state of these proteins in rat striatum was analyzed by immunoblotting 15 min and 2 h after amphetamine (AMPH)-induced behavioral sensitization. Two weeks after the last dose of 5 mg/kg, i.p. AMPH once daily for 5 days, rats were challenged with 1 mg/kg, i.p. AMPH or saline and sacrificed 15 min or 2 h later. Sensitization to AMPH-induced behavioral activity was observed in AMPH pre-treated rats after AMPH on the challenge day. Phosphorylation of all three proteins was significantly greater 15 min after AMPH in AMPH-pre-treated than in saline-pre-treated rats. Two hours after AMPH challenge in AMPH-pre-treated rats, phospho-extracellular signal-regulated kinase and phospho-cAMP response element-binding protein immunoreactivity was still significantly elevated but not after AMPH injection in saline-pre-treated rats. In contrast, phospho-Akt was down-regulated to the same extent 2 h after acute AMPH or repeated AMPH with an AMPH challenge. These data implicate differential regulation of phospho-extracellular signal-regulated kinase, phospho-cAMP response element-binding protein versus phospho-Akt in sensitized responses to AMPH. 相似文献
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Paul Mangeat Jacques Marvaldi Ounissa Ait Ahmed Guy Marchis-Mouren 《Regulatory peptides》1981,1(6):397-414
Vasoactive intestinal peptide (VIP), secretin, catecholamines and prostaglandin E1 (PGE1) in the presence of a cyclic nucleotide phosphodiesterase inhibitor stimulate the accumulation of cyclic AMP in two colorectal carcinoma cell lines (HT 29 and HRT 18) with subsequent activation of the cyclic AMP-dependent protein kinases. In HT 29 cells incubated without phosphodiesterase inhibitor, 10?9 M VIP promotes a rapid and specific activation of the low cyclic AMP phosphodiesterase (1.7-fold); at 25°C the effect is maintained for more than 15 min, while at 37°C the activity returns to basal value within 15 min. As shown by dose-response studies, VIP is by far the most effective inducer () of the cyclic AMP phosphodiesterase activity; partial activation of the enzyme is obtained by 3 · 10?7 M secretin, 10?5 M isoproterenol and 10?5 M PGE1; PGE2 and epinephrine are without effect. In HRT 18 cells VIP is less active () whereas 10?6 M PGE1, 10?6 M PGE2 and 10?5 M epinephrine are potent inducers of the phosphodiesterase activity. The positive cell response to dibutyryl-cyclic AMP further indicates that cyclic AMP is a mediator in the phosphodiesterase activation process. The incubation kinetics and dose response effects of the various agonists on the cyclic AMP-dependent protein kinase activity determined for both cell types in the same conditions show a striking similarity to those of phosphodiesterase. Thus coordinate regulation of both enzymes by cyclic AMP was observed in all incubation conditions. 相似文献
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Yasuo Totsuka Mehdi S. Ferdows Thor B. Nielsen James B. Field 《Biochimica et Biophysica Acta (BBA)/General Subjects》1983,756(3):319-327
Forskolin (40 μM) stimulated adenylate cyclase activities of bovine thyroid plasma membranes without pthe addition of guanine nucleotides. GDP had little effect on the forskolin-stimulated adenylate cyclase activity while Gpp[NH]p (0.1–1.0 μM) decreased it. In the presence of TSH (10 mU/0.11), Gpp[NH]p no longer caused inhibition. Forskolin did not affect phosphodiesterase activities of thyroid homogenates. Forskolin (10 μM) rapidly increased cAMP levels in bovine thyroid slices both in the absence and presence of a phosphodiesterase inhibitor. The effect of TSH (50 mU/ml) on cAMP levels was additive or greater than additive to that of forskolin. An initial 2-h incubation of slices with forskolin did not decrease their subsequent cAMP responses to either forskolin and/or TSH while similar treatment of slices with TSH induced desensitization of the cAMP response to TSH, but not to forskolin. Forskolin (10 μM) as well as TSH (50 mU/ml) activated cAMP-dependent protein kinase of slices in the absence of a phosphodiesterase inhibitor. Although forskolin activated the adenylate cyclase cAMP system, it did not stimulate iodide organification or glucose oxidation, effects which have been attributed to cAMP. In fact, forskolin inhibited these parameters and 32P incorporation into phospholipids as well as their stimulation by TSH. These results indicate that an increase in cAMP levels and cAMP-dependent protein kinase activity in thyroid slices may not necessarily reproduce the effects of TSH on the thyroid. 相似文献
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Ernst-Georg Krause Sabine Bartel Inge Beyerdörfer Wolfgang Freier Karl Gerber Dankwart Obst 《Molecular and cellular biochemistry》1989,89(2):181-186
Summary Oscillation of cyclic AMP and in the activity ratio of cyclic AMP-dependent protein kinase and of glycogen phosphorylase with the cardiac cycle were demonstrated in the canine heart in situ. For tissue sampling an ECG (R-wave)-triggered, automatically working push-freeze-drill apparatus was developed which allows intraventricular cryobiopsies from the left ventricular muscle of anaesthetized open-chest dogs. The nucleotide cyclic AMP oscillated with the cardiac cycle during normal working condition, the higher cyclic AMP level occuring during systole. Cyclic GMP was assayed to be without oscillatory changes during the contraction-relaxation cycle. The rise in the activity ratio of protein kinase was found to coincide with the maximum in the level of cyclic AMP. Propranolol pretreatment prevents the transient in the level of the nucleotide as well as in the activity ratio of the kinase indicating i) a causal relationship between these changes and ii) a neurohumoral, beat-to-beat regulation by catecholamines released from the sympathetic nerve endings within the heart. Contrary the activity ratio of phosphorylase retains its transient changes during the cardiac cycle in the presence of propranolol, indicating a Ca-mediated activation of phosphorlase kinase during the contraction process. 相似文献
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Characterization of a fluorescent substrate for the adenosine 3',5'-cyclic monophosphate-dependent protein kinase 总被引:1,自引:0,他引:1
A synthetic tetradecapeptide derived from the phosphorylation site of the beta-subunit of phosphorylase kinase (Arg-Thr-Lys-Arg-Ser-Gly-Ser-Val-Tyr-Glu-Pro-Leu-Lys-Ile) is a highly efficient substrate for the cAMP-dependent protein kinase, exhibiting a 36% decrease in the intrinsic tyrosine fluorescence on phosphorylation. The fluorescence changes in continuous assays were monitored to demonstrate the roles of protein kinase effectors (cAMP, the type II regulatory subunit, and the 8000-Da heat-stable inhibitor) in the regulation of the enzyme and to determine Km and Vmax. The phosphorylation reaction requires 1 mol ATP/mol peptide. Amino acid analysis demonstrates the presence of phosphoserine in the phosphorylated peptide. Auxiliary experiments show that tyrosine phosphorylation can also be detected fluorometrically and distinguished from serine or threonine phosphorylation. 相似文献
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Pawelczyk T Sakowicz M Podgorska M Szczepanska-Konkel M 《Experimental cell research》2003,286(1):152-163
The activity of adenosine kinase (AK) was significantly impaired in splenocytes isolated from diabetic rats. Administration of insulin to diabetic animals restored AK activity, protein, and mRNA levels in diabetic splenocytes. Experiments performed on cultured rat lymphocytes demonstrated that insulin did not change the stability of AK mRNA. Insulin induced AK gene expression in a dose- and time-dependent manner. Maximal increases in AK mRNA (3.9-fold) and activity level (3.7-fold) were observed at the fourth and fifth hours of cell incubation with 10 nM insulin, respectively. The insulin effect on AK expression was not influenced by dibutyryl cAMP (dcAMP). On the other hand dcAMP weakly increased (1.7-fold) basal expression of AK. Exposure of rat lymphocytes to wortmannin, an inhibitor of phosphatidylinositol 3-kinase (PI3K), or rapamycin, an inhibitor of mTOR, did not affect the ability of insulin to stimulate expression of AK. Prior treatment of the cells with 10 microM PD98059, an inhibitor of mitogen-activated protein kinase (MAPK) kinase (MEK) completely blocked insulin-stimulated expression of AK gene. Insulin produced a significant transient increase in the tyrosine phosphorylation of ERK1/2, and PD98059 inhibited this phosphorylation. Furthermore exposure of cells to insulin has resulted in transient phosphorylation of Elk-1 on Ser-383 and sustained elevation of c-Jun and c-Fos protein. The maximal phosphorylation of Elk-1 was observed at 15 min, and was blocked by PD98059. We concluded that insulin stimulates AK gene expression through a series of events occurring sequentially. This includes activation of the MAPK cascade and subsequent phosphorylation of Elk-1 followed by increased expression of c-fos and c-jun genes. 相似文献