Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

---

Query: EC:3.1.3.16 (calcineurin)
17,112 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Studies on sphingomyelin metabolism in rat hepatocytes were facilitated by the use of choline-deficient cells which allowed for the rapid labeling of phosphatidylcholine and as a result sphingomyelin. Pulse and pulse-chase studies with [methyl-3H]choline and [methyl-3H]methionine demonstrated that both compounds were effectively used for sphingomyelin biosynthesis and that newly made and pre-existing phosphatidylcholine could be used for sphingomyelin biosynthesis. When hepatocytes were incubated with brefeldin A, there was a 2.4-fold stimulation of the conversion of phosphatidylcholine into sphingomyelin. Since brefeldin A causes collapse of the cis/medial Golgi into the endoplasmic reticulum the stimulation of sphingomyelin biosynthesis could be due to more rapid access of the labeled phosphatidylcholine in the endoplasmic reticulum to sphingomyelin synthase in the collapsed Golgi. Forskolin inhibited the brefeldin A-induced stimulation of sphingomyelin biosynthesis. To investigate whether or not phosphorylation reactions regulate sphingomyelin metabolism, hepatocytes were incubated with okadaic acid, a potent inhibitor of protein phosphatases 1 and 2A. Rather than stimulating sphingomyelin biosynthesis, okadaic acid enhanced the catabolism of sphingomyelin. In contrast, a cyclic AMP analogue and forskolin had no effect on sphingomyelin biosynthesis or catabolism. Surprisingly, other pulse-chase studies demonstrated that okadaic acid stimulated the catabolism of only newly made sphingomyelin. The brefeldin A and okadaic acid effects were independent of lysosomal involvement. Subcellular fractionation studies revealed that brefeldin A and okadaic acid effects were generalized in all sphingomyelin containing membranes. The brefeldin A studies suggest that the rate of transfer of phosphatidylcholine from the endoplasmic reticulum to the Golgi might be limiting for sphingomyelin biosynthesis. The okadaic acid studies indicate that the catabolism of sphingomyelin by a sphingomyelinase is regulated by an unidentified protein kinase and by either protein phosphatase 1 and/or 2A activity in hepatocytes.
...
PMID:Stimulation of sphingomyelin biosynthesis by brefeldin A and sphingomyelin breakdown by okadaic acid treatment of rat hepatocytes. 161 52

We have studied cAMP-dependent phosphorylation of sodium channels in rat brain neurons maintained in primary culture. In back phosphorylation studies, cells were treated with drugs to increase intracellular cAMP and sodium channels were solubilized and isolated by immunoprecipitation. Surface and intracellular pools of sodium channels were isolated separately. Purified channels were then phosphorylated with [gamma-32P]ATP by the catalytic subunit of cAMP-dependent protein kinase to incorporate 32P into available cAMP-dependent phosphorylation sites. The amount of 32P incorporated in vitro is inversely proportional to the extent of endogenous phosphorylation. Incubation of cells with forskolin (0.1-100 microM), 8-Br-cAMP (0.1-10 mM), or isobutylmethylxanthine (0.01-1.0 mM) inhibited subsequent incorporation of 32P into isolated sodium channels by 70-80%, indicating that treatment of cells with these drugs had increased endogenous phosphorylation to nearly maximum levels. The phosphopeptides phosphorylated in vivo and in vitro were identical. To examine the magnitude of basal phosphorylation and the extent of stimulated phosphorylation, the amount of 32P incorporated into sodium channels from control and stimulated cells was compared to that from matched samples which had been dephosphorylated with calcineurin. Sodium channels from control cells incorporated approximately 2-fold more 32P after dephosphorylation, indicating that cAMP-dependent sites on the channel are at least 47% phosphorylated in the basal state. Sodium channels from forskolin-treated cells incorporated 7-8-fold more 32P after dephosphorylation, indicating that cAMP-dependent phosphorylation sites are 80-90% phosphorylated after stimulation. Cell surface and intracellular pools of sodium channels were phosphorylated similarly. In cells metabolically labeled with 32P, cell surface sodium channels incorporated 2.7 mol of phosphate/mol of channel. Forskolin stimulated 32P incorporation into sodium channels 1.3-fold, consistent with the results obtained by back phosphorylation. We conclude that the rat brain sodium channel is substantially phosphorylated in both the cell surface and intracellular pools in vivo in unstimulated rat brain neurons, and the extent of phosphorylation is increased to 80-90% of maximum phosphorylation by agents that elevate intracellular cAMP.
...
PMID:Cyclic-AMP-dependent phosphorylation of voltage-sensitive sodium channels in primary cultures of rat brain neurons. 244 66

We have identified and studied a posttranscriptional mechanism of lactate dehydrogenase A (LDH) subunit gene expression at the level of mRNA stability. Using the well differentiated rat C6 glioma cell line as a model system, the effects of activators of the protein kinase A and C pathways on the half-life of LDH A mRNA were measured by two independent methods: 1) by the RNA synthesis inhibitor-chase method using actinomycin D, and 2) by analysis of decay of LDH A [3H]mRNA in [3H]uridine-labeled cells. By each method, the half-life of relatively short-lived LDH A mRNA was increased 5- to 7-fold in 8- (4-chloro-phenylthio) cAMP or forskolin-treated and about 3-fold in 12-0-tetradecanoylphorbol-13- acetate (TPA) or dioctanoylglycerol-treated cells. Forskolin acted synergistically with TPA to prolong LDH A mRNA half-life from 55 min to more than 20 h. The relatively rapid basal decay rate of LDH A mRNA was also considerably slowed in the presence of the protein phosphatase inhibitor okadaic acid, suggesting a functional role for protein phosphorylation in the stabilization process. In glioma cells stably transformed with a protein kinase A catalytic subunit expression vector, overexpression of the catalytic subunit stabilized LDH mRNA to the degree seen in forskolin-treated cells. In cells transfected with a protein kinase A inhibitor-expression vector, cAMP-mediated stabilization of LDH A mRNA half-life was prevented. Furthermore, both staurosporin and 3- [1-(3-dimethylaminopropyl)-indol-3-yl]-3-(indol- 3-yl)- maleimide, inhibitors of protein kinase C, prevented the TPA-induced stabilization of LDH A mRNA. We conclude from the experimental data that the protein kinase A and C signal pathways play an active functional role in regulating LDH A mRNA stability and act cooperatively to achieve LDH A mRNA stability regulation.
...
PMID:Lactate dehydrogenase A subunit messenger RNA stability is synergistically regulated via the protein kinase A and C signal transduction pathways. 747 96

Intracellular recordings were obtained from myenteric AH neurons of guinea pig ileum in vitro. Slow excitatory synaptic responses associated with decreased potassium conductance (gK), inhibition of the spike afterhyperpolarization current (AHC), and increased chloride conductance (gCl) were mimicked by senktide, a neurokinin3 receptor agonist. Intracellular guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) decreased gK and increased gCl irreversibly after nerve stimulation or senktide application. Myenteric neurons in pertussis toxin (PTX)-treated tissues responded normally to senktide and nerve stimulation. Forskolin and phorbol 12,13-dibutyrate (PDBu) inhibited gK and the AHC but did not activate gCl. The AHC was not reduced by subthreshold concentrations of forskolin (10 nM) or PDBu (3 nM) alone but was inhibited by forskolin and PDBu applied together. Inhibitors of phospholipase C (D-609) or protein kinases (staurosporine) reduced slow synaptic and senktide responses. The protein phosphatase inhibitor, calyculin A, caused an inward current, a decrease in gK, and AHC inhibition but did not activate gCl. We conclude that slow excitatory synaptic responses are mediated by PTX-insensitive G proteins and activation of phospholipase C and protein kinases. Forskolin and PDBu activate pathways that inhibit gK. The mechanisms for activation of gCl are unknown.
...
PMID:Signal-transduction pathways causing slow synaptic excitation in guinea pig myenteric AH neurons. 749 63

The characteristics of ATP release evoked by forskolin and ouabain from atrial segments of guinea-pig were evaluated under electrical stimulation. Forskolin (1 microM) produced a massive release of ATP together with a positive inotropic response. Both 30 microM W-7 (N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide.HCI), a calmodulin antagonist, and 30 microM vinblastine, a mitotic inhibitor, markedly inhibited the evoked release of ATP without affecting the evoked contraction. However, 100 microM N-ethylmaleimide abolished completely the basal and drug-evoked ATP release and further the evoked contraction. Both the ATP release and contraction evoked by ouabain (3 microM) were similarly affected by W-7, vinblastine and n-ethylmaleimide. The release of ATP, but not the contraction, evoked by forskolin was strongly suppressed by 10 microM okadaic acid, a protein phosphatase inhibitor. The suppression by okadaic acid of the evoked release was thoroughly antagonized in the presence of 0.01 microM PMA (phorbol 12-myristate 13-acetate), but not 10 microM H-7 (1-(5-isoquinolinesulfonyl)-2-methylpiperazine). These results suggest that forskolin, like ouabain, may dominantly cause the neuronal release of ATP from cardiac adrenergic nerves, although the possible participation of release from muscular sources cannot be ignored.
...
PMID:Possible neuronal origin of ATP release evoked by forskolin and ouabain from guinea-pig atrial segments. 749 79

We have stably introduced expression vectors for the glucocorticoid receptor and a sensitive, hormone-responsive reporter (mouse mammary tumor virus-luciferase) into a human breast carcinoma-derived cell line. Employing this cell line, we have conducted a detailed examination of the induction of glucocorticoid-regulated genes and the phosphorylation of glucocorticoid receptor following pharmacologic manipulation of cell signaling pathways. The hormone response can be enhanced from 2 to 10-fold by activators of protein kinase A, protein kinase C, and inhibitors of protein phosphatase. Forskolin and 8-bromoadenosine 3':5'-cyclic monophosphate (BrcAMP), but not BrcGMP, enhance the hormone effect, yet surprisingly, phosphodiesterase inhibitors, isobutylmethylxanthine and Ro20-1724, strongly inhibit hormone-mediated induction of the reporter gene. These treatments do not alter cellular receptor content, dexamethasone binding, nor hormone-mediated receptor down-regulation. Tryptic peptide analysis of 32P-labeled receptor reveals that neither BrcAMP, isobutylmethylxanthine, nor the tumor promoter and protein kinase C activator, 12-O-tetradecanoyl-phorbol-13-acetate, detectably alter the state of glucocorticoid receptor phosphorylation. The only agent which alters receptor phosphorylation is the protein phosphatase inhibitor okadaic acid, but only at concentrations higher than required for maximum effects on glucocorticoid receptor transactivation. We propose that these effectors do not modify receptor directly but alter its interaction with transcription complexes.
...
PMID:Modulation of cell signaling pathways can enhance or impair glucocorticoid-induced gene expression without altering the state of receptor phosphorylation. 769 81

Tau is a neuron-specific, microtubule-associated protein that forms paired helical filaments (PHFs) of Alzheimer's disease when aberrantly phosphorylated. We have attempted to elucidate the protein kinases and phosphatases that regulate tau phosphorylation. Incubation of rat, human, and rhesus monkey temporal neocortex slices with the phosphatase inhibitor okadaic acid induced epitopes of tau similar to those found in PHFs. Okadaic acid (1-20 microM) induced variant forms of tau at 60-68 kDa, which were recognized by the monoclonal antibodies Alz-50 (in humans only) and 5E2 and two polyclonal antipeptide antisera, OK-1 and OK-2. The phosphorylation-sensitive monoclonal antibody Tau-1 failed to recognize the slowest mobility forms of tau after okadaic acid treatment. FK-520 (1-10 microM), a potent inhibitor of calcineurin activity, was tested in brain slices and found not to alter tau mobility. However, combinations of FK-520 (5 microM) and okadaic acid (100 nM) caused tau mobility shifts similar to those seen after 10 microM okadaic acid treatment; similar results were seen using the calcineurin-selective inhibitor cypermethrin. Treatment of human slices with 10 microM okadaic acid decreased both protein phosphatase 2A and calcineurin activity; FK-520 inhibited only protein phosphatase 2B activity. A proposed tau-directed kinase, 42-kDa mitogen-activated protein kinase (p42mapk), was activated by okadaic acid (> 100 nM) but not FK-520 (5 microM). Nerve growth factor (100 ng/ml) activated p42mapk, particularly when used in combination with 100 nM okadaic acid; changes in tau mobility were seen when this kinase was activated. Forskolin (2 microM) antagonized the effects of nerve growth factor on both p42mapk activity and tau phosphorylation; forskolin alone had little effect on PHF-like tau formation induced by phosphatase inhibitors. These results outline complex interactions between tau-directed protein kinases and protein phosphatases and suggest potential sites for therapeutic intervention.
...
PMID:Tau phosphorylation in brain slices: pharmacological evidence for convergent effects of protein phosphatases on tau and mitogen-activated protein kinase. 772 35

Treatment of striatal synaptosomes with the protein phosphatase inhibitor okadaic acid significantly decreased gamma-aminobutyric acid (GABA) uptake, indicating that the GABA transporter may be regulated by phosphorylation. Forskolin and 8-bromoadenosine-3,5-cyclic monophosphate (8-br-cAMP) inhibited GABA uptake to the same extent as okadaic acid, suggesting the involvement of protein kinase A in GABA transporter regulation. In contrast, the same treatments did not alter dopamine (DA) uptake into striatal synaptosomal preparations. The results suggest that the structurally related GABA and DA transporters may be subject to different post-translational regulation.
...
PMID:Dopamine and gamma-aminobutyric acid transporters: differential regulation by agents that promote phosphorylation. 793 2

To elucidate the mechanism causing the transient accumulation of intracellular cAMP in the FRTL-5 thyroid cell line, the short-term effect of thyroid-stimulating hormone (TSH) on phosphodiesterase (PDE) activity was studied. Together with an increase in cAMP levels, TSH produced a significant increase in total PDE activity as early as 3 min, with a maximal stimulation reached after 15 min. This short-term increase in PDE activity was dependent on the TSH concentration (ED50 = 4 x 10(-11) M TSH). Forskolin and dibutyryl cAMP produced an even larger stimulation than that produced by TSH, suggesting that the effect of TSH is mediated by cAMP. To determine the properties of the PDE forms activated by TSH, antibodies specific for the cAMP-PDEs were used to immunoprecipitate the PDEs present in control cells, and cells incubated for 15 min in the presence of 10 nM TSH. Comparison of the activity recovered in the immunoprecipitation pellets demonstrated that TSH produced more than a 2.5-fold increase in the cAMP-PDE form(s) recognized by this antibody. Conversely, the activity remaining in the supernatants was not affected by the TSH treatment. Most of the activity recovered in the immunoprecipitation pellets (90%) was inhibited by 10 microM Rolipram, an inhibitor specific for the high affinity cAMP-PDEs. No TSH stimulation of the Rolipram-insensitive PDE activity could be observed under these conditions. Western blot analyses with two different cAMP-PDE specific antibodies showed that a 15-min stimulation with TSH induced the appearance of a new band with electrophoretic mobility slower than the polypeptide present in unstimulated cells. The appearance of this band did not require ongoing protein synthesis because it occurred in the presence of cycloheximide. Metabolic [32P]orthophosphate labeling of intact FRTL-5 cells indicated that the TSH treatment caused an increased 32P incorporation into a polypeptide that co-purified with the stimulated PDE activity and had an electrophoretic mobility identical to that of the cAMP-PDE. Okadaic acid, a potent inhibitor of protein phosphatase 1 and protein phosphatase 2A, elicited a potentiation of the TSH-stimulated PDE activity. The stimulating of a PDE with the same immunological properties and Rolipram sensitivity as the cAMP-PDE stimulated by TSH in the intact cells was reproduced, in a cell-free system, by incubating soluble extracts from FRTL-5 cells with the catalytic subunit of cAMP-dependent protein kinase. These data provide evidence that TSH produces a rapid activation of a cAMP-PDE in the FRTL-5 cells through a cAMP-dependent phosphorylation.
...
PMID:The short-term activation of a rolipram-sensitive, cAMP-specific phosphodiesterase by thyroid-stimulating hormone in thyroid FRTL-5 cells is mediated by a cAMP-dependent phosphorylation. 813 62

We have previously shown [B. Illek, H. Fischer, G. F. Santos, J. H. Widdicombe, T. E. Machen, and W. W. Reenstra, Am. J. Physiol. 268 (Cell Physiol. 37): C886-C893, 1995] that genistein, a tyrosine kinase inhibitor, activates the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel in NIH/3T3 cells that have been stably transfected with an expression vector for the CFTR (NIH-CFTR cells). In this study, we present evidence suggesting that both genistein and the serine/threonine protein phosphatase (PPase) inhibitor calyculin A activate the CFTR by inhibiting PPase activity. As measured by 125I efflux, genistein and calyculin A stimulate the CFTR to approximately 50% of the maximal activity with forskolin. Neither agonist increases CFTR activity at saturating forskolin concentrations, but genistein and calyculin A have an additive effect on CFTR activity. Forskolin, but neither genistein nor calyculin A, stimulates protein kinase A(PKA) activity. The PKA inhibitor H-89 inhibits CFTR activation and in vivo phosphorylation by all three agonists. Proteolytic digestion of in vivo phosphorylated CFTR suggests that the CFTR is phosphorylated on the same sites during stimulation with genistein and forskolin but on different sites stimulation with calyculin A. The data suggest that genistein and calyculin A inhibit different PPase activities, allowing CFTR phosphorylation and partial stimulation, by a basal PKA activity.
...
PMID:CFTR chloride channel activation by genistein: the role of serine/threonine protein phosphatases. 877 6


1 2 3 Next >>

---