Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.3.16 (calcineurin)
 17,112 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In summary, we propose that acute ammonia intoxication leads to increased extracellular concentration of glutamate in brain and results in activation of the NMDA receptor. Activation of this receptor mediates ATP depletion and ammonia toxicity since blocking the NMDA receptor with MK-801 prevents both phenomena. Ammonia-induced metabolic alterations (in glycogen, glucose, pyruvate, lactate, glutamine, glutamate, etc) are not prevented by MK-801 and, therefore, it seems that they do not play a direct role in ammonia-induced ATP depletion nor in the molecular mechanism of acute ammonia toxicity. The above results suggest that ammonia-induced ATP depletion is due to activation of Na+/K(+)-ATPase, which, in turn, is a consequence of decreased phosphorylation by protein kinase C. This can be due to decreased activity of PKC or to increased activity of a protein phosphatase. We also show that L-carnitine prevents glutamate toxicity in primary neuronal cultures. The results shown indicate that carnitine increases the affinity of glutamate for the quisqualate type (including metabotropic) of glutamate receptors. Also, blocking the metabotropic receptor with AP-3 prevents the protective effect of L-carnitine, indicating that activation of this receptor mediates the protective effect of carnitine. We suggest that the protective effect of carnitine against acute ammonia toxicity in animals is due to the protection against glutamate neurotoxicity according to the above mechanisms.
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PMID:Molecular mechanism of acute ammonia toxicity and of its prevention by L-carnitine. 774 Oct 17

DNA topoisomerase I was partially purified from the hepatopancreas of the shrimp Penaeus japonicus. The specific activity of the final preparation was 7,000,000 units/mg of protein with SV40 viral DNA as substrate. SDD-polyacrylamide gel electrophoresis of the final preparation yielded two major bands of proteins with M(r) 70,000 and M(r) 67,000, as well as less intense bands of proteins with M, 64,000 and M(r) 56,000. Incubation of the partially purified enzyme fraction with rabbit antiserum against human DNA topoisomerase I, allowed all these proteins except that of M(r) 56,000, to be positively reacted. Treatment of the partially purified DNA topoisomerase I with tyrosine kinase p43v-abl resulted in phosphorylation of only the two major subunits. Phosphorylation by tyrosine kinase p43v-abl or dephosphorylation by phosphotyrosyl protein phosphatase resulted in a decrease of the enzymatic activity. The treatment with shrimp alkaline phosphatase abolished the enzymatic activity of the purified DNA topoisomerase I in a dose-dependent manner. Thus, the DNA topoisomerase I was apparently isolated from the hepatopancreas of the shrimp P. japonicus in a phosphorylated form, and this phosphorylation was essential for expression of enzymatic activity in vitro. The activity of DNA topoisomerase I is inhibited by ZnCl2, CuCl2 and Pb(NH3)3 at millimolar concentrations, but less inhibition was observed with CaCl2.
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PMID:Modification of DNA topoisomerase I enzymatic activity with phosphotyrosyl protein phosphatase and alkaline phosphatase from the hepatopancreas of the shrimp Penaeus japonicus (Crustacea:Decapoda). 875 89

As a first step towards isolation of autophagic sequestering membranes (phagophores), we have purified autophagosomes from rat hepatocytes. Lysosomes were selectively destroyed by osmotic rupture, achieved by incubation of hepatocyte homogenates with the cathepsin C substrate glycyl-phenylalanyl-naphthylamide (GPN). Mitochondria and peroxisomes were removed by Nycodenz gradient centrifugation, and cytosol, microsomes and other organelles by rate sedimentation through metrizamide cushions. The purified autophagosomes were bordered by dual or multiple concentric membranes, suggesting that autophagic sequestration might be performed either by single autophagic cisternae or by cisternal stacks. Okadaic acid, a protein phosphatase inhibitor, disrupted the hepatocytic cytokeratin network and inhibited autophagy completely in intact hepatocytes, perhaps suggesting that autophagy might be dependent on intact intermediate filaments. Vinblastine and cytochalasin D, which specifically disrupted microtubules and microfilaments, respectively, had relatively little (25-30%) inhibitory effect on autophagic sequestration. In a cryo-ultrastructural study, the various autophagic-lysosomal vacuoles were immunogold-labelled, using the cytosolic enzyme superoxide dismutase as an autophagic marker, Lgp120 as a lysosomal membrane marker, and bovine serum albumin as an endocytic marker. Vinblastine (50 microM) was found to inhibit both autophagic and endocytic flux into the lysosomes, with a consequent reduction in lysosomal size. Asparagine (20 mM) caused swelling of the lysosomes, probably as a result of the ammonia formation that could be observed at this high asparagine concentration. Autophagosomes and amphisomes (autophagic-endocytic, prelysosomal vacuoles) accumulated in asparagine-treated cells, reflecting an inhibition of autophagic flux that might be a consequence of lysosomal dysfunction.
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PMID:Structural aspects of autophagy. 886 Sep 99

We proposed that acute ammonia toxicity is mediated by activation of NMDA receptors. To confirm this hypothesis we have tested whether different NMDA receptor antagonists, acting on different sites of NMDA receptors, prevent death of mice induced by injection of 14 mmol/Kg of ammonium acetate, a dose that induces death of 95% of mice. MK-801, phencyclidine and ketamine, which block the ion channel of NMDA receptors, prevent death of at least 75% of mice. CPP, AP-5, CGS 19755, and CGP 40116, competitive antagonists acting on the binding site for NMDA, also prevent death of at least 75% of mice. Butanol, ethanol and methanol which block NMDA receptors, also prevent death of mice. There is an excellent correlation between the EC50 for preventing ammonia-induced death and the IC50 for inhibiting NMDA-induced currents. Acute ammonia toxicity is not prevented by antagonists of kainate/AMPA receptors, of muscarinic or nicotinic acetylcholine receptors or of GABA receptors. Inhibitors of nitric oxide synthase afford partial protection against ammonia toxicity while inhibitors of calcineurin, of glutamine synthetase or antioxidants did not prevent ammonia-induced death of mice. These results strongly support the idea that acute ammonia toxicity is mediated by activation of NMDA receptors.
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PMID:NMDA receptor antagonists prevent acute ammonia toxicity in mice. 892 86

Ammonia is a main factor in the pathogenesis of hepatic encephalopathy. We found that acute ammonia toxicity is mediated by activation of NMDA receptors. Chronic moderate hyperammonemia prevents acute ammonia toxicity in rats. Chronic exposure of cultured neurons to 1 mM ammonia leads to impaired response of the NMDA receptor to activation by its agonists (due to decreased protein kinase C-mediated phosphorylation) and prevents glutamate (Glu) neurotoxicity. Compounds that prevent ammonia toxicity in mice (e.g. carnitine) also prevent Glu toxicity in cultured neurons. These compounds did not prevent activation of NMDA receptor or the rise of Ca2+. They interfered with subsequent steps in the toxic process. The protective effect of carnitine is mediated by activation of metabotropic Glu receptors. Agonists of mGluRs, especially of mGluR5, prevent Glu toxicity. Agonists of muscarinic receptors also prevent Glu toxicity and there seems to be an interplay between muscarinic and metabotropic Glu receptors in the protective effect. We have tried to identify intracellular events involved in the process of neuronal death. It is known that the rise of Ca2+ is an essential step. Glu leads to depletion of ATP; some compounds (e.g. carnitine) prevent Glu-induced neuronal death without preventing ATP depletion: additional events are required for neuronal death. Glu induces activation of Na+/K+-ATPase, which could be involved in the toxic process. Inhibitors of protein kinase C, calcineurin or nitric oxide synthase prevent Glu toxicity. Our results indicate that Glu toxicity can be prevented at different steps or by activating receptors coupled to the transduction pathways interfering with the toxic process. Agents acting on these steps could prevent excitotoxicity in vivo in animals.
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PMID:Neurotoxicity of ammonia and glutamate: molecular mechanisms and prevention. 974 28

As a substitute for M(H2O)2+6, Co(NH3)3+6 was found to activate calcineurin with para-nitrophenyl phosphate as substrate. Kinetics for calcineurin catalyzed hydrolysis of para-nitrophenyl phosphate at pH 7.0 with Mn2+, Mg2+, Co2+, and Co(NH3)3+6 were compared. Although kcat and Km were different with the metals, values of kcat/Km were nearly identical for Mn2+ and Mg2+, but lower for Co2+ and Co(NH3)3+6. The concentration of each metal providing half-maximal activation, designated Kact, was evaluated as 15.9 mM for Co(NH3)3+6, compared to Kact = 0.17 mM for Mn2+ and Co2+ and 6.3 mM for Mg2+, respectively. Comparing kcat/Kcat showed that Co(NH3)3+6 was a 170-fold poorer activator of calcineurin than was Mn2+, but only 1.5-fold poorer than Mg2+. Activation by Co(NH3)3+6 indicated that activation of calcineurin by exogenous metal ions can proceed via an outer coordination sphere reaction mechanism with no requirement for the direct coordination of substrate by metal. Because Co(NH3)3+6 was found to support calcineurin activity, the related compound [Co-(ethylenediamine)3]3+ (or Co(en)3+3) was tested as a possible activator. Co(en)3+3 did not support calcineurin activity but did inhibit calcineurin. Co(en)3+3 showed competitive inhibition kinetics with either Mn2+ or pNPP as the varied ligand and the other at a fixed, subsaturating concentration. Inorganic phosphate was used as a known competitive inhibitor to pNPP (B. L. Martin and D. J. Graves, J. Biol. Chem. 261, 14545-14550, 1986) and showed uncompetitive inhibition with Mn2+ as the varied ligand. These patterns are consistent with the mechanism of ligand binding to calcineurin being ordered with metal preceding substrate. Prior formation of a metal-substrate complex was not required for association with calcineurin.
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PMID:Effect of substitution inert metal complexes on calcineurin. 1033 77

Hyperammonemia is considered the main cause for the neurological alterations found in hepatic failure. However, the mechanisms by which high ammonia levels impair cerebral function are not well understood. It has been shown that chronic hyperammonemia impairs signal transduction pathways associated with NMDA receptors and also alters phosphorylation of some neuronal proteins. The aim of the present work was to analyze the effects of chronic exposure to ammonia on phosphorylation of microtubule-associated protein 2 (MAP-2) in intact neurons in culture and to assess whether modulation of MAP-2 phosphorylation by glutamate receptor-associated transduction pathways is altered in neurons chronically exposed to ammonia. It is shown that chronic exposure to ammonia increases basal phosphorylation of MAP-2 by approximately 70%. This effect seems to be due to a decreased tonic activation of NMDA receptors and of calcineurin. Chronic exposure to ammonia also alters the modulation of MAP-2 phosphorylation by NMDA receptors and metabotropic glutamate receptors. In neurons exposed to ammonia, treatment with NMDA for 30 min induced a significant decrease in phosphorylation of MAP-2. Activation of metabotropic glutamate receptors with (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid significantly increased phosphorylation of MAP-2 in control neurons, whereas in neurons exposed to ammonia the response was the opposite, with 1-aminocyclopentane-1,3-dicarboxylic acid inducing a dephosphorylation of MAP-2. These results indicate that ammonia alters significantly signal transduction pathways associated with different types of glutamate receptors. This would lead therefore to significant alterations in glutamatergic neurotransmission, which would contribute to the neurological alterations found in hyperammonemia and in hepatic encephalopathy.
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PMID:Chronic exposure to ammonia alters pathways modulating phosphorylation of microtubule-associated protein 2 in cerebellar neurons in culture. 1058 18

Acute administration of large doses of ammonia leads to the rapid death of animals. This article reviews the role of excessive activation of N-methyl-D-aspartate (NMDA) receptors in the mediation of ammonia-induced mortality. The studies reviewed here show that acute intoxication with large doses of ammonia leads to the activation of NMDA receptors in brain in vivo. Moreover, excessive activation of NMDA receptors is responsible for ammonia-induced death of animals, which is prevented by different antagonists of NMDA receptors. This article also reviews the studies showing that activation of NMDA receptors is also responsible for the following effects of acute ammonia intoxication: (1) depletion of brain ATP, which, in turn, leads to release of glutamate; (2) activation of calcineurin and dephosphorylation and activation of Na+/K+-ATPase in brain, thus increasing ATP consumption; (3) impairment of mitochondrial function and calcium homeostasis at different levels, thus decreasing ATP synthesis; (4) activation of calpain that degrades the microtubule-associated protein MAP-2, thus altering the microtubular network; (5) increased formation of nitric oxide (NO) formation, which, in turn, reduces the activity of glutamine synthetase, thus reducing the elimination of ammonia in brain.
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PMID:Molecular mechanism of acute ammonia toxicity: role of NMDA receptors. 1202 Jun 9

We previously reported that ammonia induced apoptosis in cultured rat hippocampal neurons with moderate increases in the intracellular calcium concentration and decreases in phospho-BAD levels. Since this suggested the involvement of calcineurin in the apoptosis, the effects of calcineurin inhibitors, 1 microM cyclosporin A and 1 microM FK506, on the ammonia-induced neuronal apoptosis were tested. Both of the inhibitors abolished the neuronal apoptosis assessed by double staining with Hoechst 33258 and anti-neurofilament antibody, and the ammonia-induced decrease in phospho-BAD Ser(155) level. Thus, calcineurin appeared to be involved in the dephosphorylation of BAD at the sites including Ser(155) in ammonia-induced apoptosis.
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PMID:Calcineurin-mediated BAD Ser155 dephosphorylation in ammonia-induced apoptosis of cultured rat hippocampal neurons. 1503 16

To identify the genes involved in the partial resistance of sunflower (Helianthus annuus) to the necrotrophic fungus Phoma macdonaldii, we developed a 1000-element cDNA microarray containing carefully chosen genes putatively involved in primary metabolic pathways, signal transduction and biotic stress responses. A two-pass general linear model was used to normalize the data and then to detect differentially expressed genes. This method allowed us to identify 38 genes differentially expressed among genotypes, treatments and times, mainly belonging to plant defense, signaling pathways and amino acid metabolism. Based on a set of genes whose differential expression was highly significant, we propose a model in which negative regulation of a dual-specificity MAPK phosphatase could be implicated in sunflower defense mechanisms against the pathogen. The resulting activation of the MAP kinase cascade could subsequently trigger defense responses (e.g. thaumatin biosynthesis and phenylalanine ammonia lyase activation), under the control of transcription factors belonging to MYB and WRKY families. Concurrently, the activation of protein phosphatase 2A (PP2A), which is implicated in cell death inhibition, could limit pathogen development. The results reported here provide a valuable first step towards the understanding and analysis of the P. macdonaldii-sunflower interaction.
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PMID:A cDNA microarray approach to decipher sunflower (Helianthus annuus) responses to the necrotrophic fungus Phoma macdonaldii. 1662 74


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