Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.13 (protein kinase C)
 49,245 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In addition to a role for de novo protein synthesis in apoptosis we have previously shown that activation of a protein phosphatase or loss of activity of a kinase is also important in radiation-induced apoptosis in human cells [Baxter, and Lavin (1992): J Immunol 148:149-1954]. We show here that some inhibitors of protein kinases exacerbate radiation-induced apoptosis in the human cell line BM13674. The specific protein kinase A inhibitor isoquinoline sulfonamide (20 microM) gave rise to significantly increased levels of apoptosis at 2-6 h postirradiation compared to values after radiation exposure only. The same concentration of isoquinolinesulfonamide, which was effective in increasing apoptosis, reduced activity markedly. A 66% inhibition of cyclic AMP-dependent protein kinase A activity occurred in unirradiated cells at this concentration of H89 and activity was reduced to 58% in irradiated cells. Calphostin C, a specific inhibitor of protein kinase C, at a concentration of 0.1 microM, which caused 68% inhibition of enzyme activity in irradiated cells, failed to enhance the level of radiation-induced apoptosis. Other kinase inhibitors did not lead to an additional increase in apoptosis over and above that observed after irradiation. The results obtained here provide further support for an important role for modification of existing proteins during radiation-induced apoptosis.
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PMID:Protein kinase A inhibitors enhance radiation-induced apoptosis. 753 51

Nerve terminals ("synaptosomes") isolated from rat brain hippocampus were loaded with the fluorescent Ca2+ indicator fura-2 and were subjected to depolarization with an elevated K+ concentration in a stopped-flow spectrophotometer to measure the activity of voltage-gated Ca2+ channels in the presynaptic membrane. Three components of Ca2+ influx were seen, which were tentatively identified as two classes of voltage-dependent Ca2+ channels with different inactivation kinetics (tau of approximately 60 ms and 1 s, respectively) and Na+/Ca2+ exchange working in the "reverse" mode. The activity of both classes of voltage-dependent Ca2+ channels was slightly augmented by the phorbol ester phorbol 12-myristate 13-acetate (PMA), an activator of protein kinase C (PKC), but the effect of PMA was markedly enhanced by the protein phosphatase inhibitor okadaic acid (OKA). The PKC inhibitors calphostin C and dihydrosphingosine (DHS) caused a prompt decrease in voltage-dependent Ca2+ channel activity, but the effect of DHS could be slowed by coaddition of OKA. These results suggest that the activity of presynaptic voltage-dependent Ca2+ channels in the hippocampus is under a dynamic balance between PKC phosphorylation (leading to activation) and protein phosphatase dephosphorylation (leading to inactivation) and that both of these metabolic pathways are tonically active in the nerve terminals.
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PMID:Protein kinase C modulates calcium channels in isolated presynaptic nerve terminals of rat hippocampus. 753 6

In nucleus basalis neurons, substance P (SP) causes a slow excitation, mediated through a pertussis toxin-insensitive G protein, by suppressing an inward rectifier K+ channel. Here we report that SP applied outside the patch pipette inhibited the single-channel activity, recorded on-cell, of the inward rectifier. The PKC inhibitors staurosporine and PKC(19-36) suppressed this effect in whole-cell mode and in on-cell single-channel mode. A diacylglycerol analog mimicked the SP effect, and PKC(19-36) suppressed this analog effect. SP irreversibly suppressed the inward rectifier in neurons treated with okadaic acid. These results indicate that a diffusible messenger mediates the SP effect, that its signal transduction involves phosphorylation by PKC, and that dephosphorylation by a serine/threonine protein phosphatase mediates its recovery.
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PMID:Protein kinase C-mediated inhibition of an inward rectifier potassium channel by substance P in nucleus basalis neurons. 753 11

To learn whether autophagy might be dependent on any of the major cytoskeletal elements, the effect of various cytoskeleton inhibitors on autophagy and cytoskeletal organization was studied in isolated rat hepatocytes. Autophagy, measured as the sequestration of endogenous lactate dehydrogenase, was completely inhibited in isolated rat hepatocytes by the protein phosphatase inhibitor okadaic acid (30 nM). Only small effects were seen with vinblastine (10 microM) or cytochalasin D (10 microM). Indirect immunofluorescence microscopy with antibody to a 55-kDa cytokeratin, corresponding to human cytokeratin 8 (CK8), revealed that whereas control cells contained a well-organized network of cytokeratin intermediate filaments, okadaic acid disrupted this network into small spherical aggregates. Treatment with cytochalasin D or vinblastine, which disrupt microfilaments and microtubules, respectively, had no detectable effect on the cytokeratin filament distribution. Neither the microtubule network (detected by indirect immunofluorescence with antibodies against alpha- and beta-tubulin) nor the actin microfilament network (detected by rhodamine-palloidin) was disrupted by okadaic acid. Naringin (100 microM), a putative protein kinase-inhibitory flavonoid, offered complete protection against the autophagy-inhibitory and cytokeratin-disruptive effects of okadaic acid. Two other flavonoids, genistein (100 microM) and prunin (100 microM), as well as KN-62 (10 microM), a specific inhibitor of Ca2+/calmodulin-dependent kinase II), likewise displayed a good ability to protect against the effect of okadaic acid upon cytokeratin organization, while no such protection was seen with H-89 (20 microM), an inhibitor of the cyclic nucleotide-dependent protein kinases, or with H-7 (100 microM), which in addition inhibits protein kinase C. The results suggest that the cytokeratin cytoskeleton of hepatocytes is subject to rapid control by phosphorylation and dephosphorylation and that cytokeratin filaments may somehow be involved in the autophagic process.
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PMID:Disruption of the cytokeratin cytoskeleton and inhibition of hepatocytic autophagy by okadaic acid. 754 Sep 86

Calcineurin is a conserved Ca2+/calmodulin-dependent protein phosphatase that plays a critical role in Ca(2+)-mediated signaling in many cells. Yeast cells lacking functional calcineurin (cna1 cna2 or cnb1 mutants) display growth defects under specific environmental conditions, for example, in the presence of high concentrations of Na+, Li+, Mn2+, or OH- but are indistinguishable from wild-type cells under standard culture conditions. To characterize regulatory pathways that may overlap with calcineurin, we performed a synthetic lethal screen to identify mutants that require calcineurin on standard growth media. The characterization of one such mutant, cnd1-8, is presented. The CND1 gene was cloned, and sequence analysis predicts that it encodes a novel protein 1,876 amino acids in length with multiple membrane-spanning domains. CND1 is identical to the gene identified previously as FKS1, ETG1, and CWH53, cnd1 mutants are sensitive to FK506 and cyclosporin A and exhibit slow growth that is improved by the addition of osmotic stabilizing agents. This osmotic agent-remedial growth defect and microscopic evidence of spontaneous cell lysis in cnd1 cultures suggest that cell integrity is compromised in these mutants. Mutations in the genes for yeast protein kinase C (pkc1) and a MAP kinase (mpk1/slt2) disrupt a Ca(2+)-dependent signaling pathway required to maintain a normal cell wall and cell integrity. We show that pkc1 and mpk1/slt2 growth defects are more severe in the absence of calcineurin function and less severe in the presence of a constitutively active form of calcineurin. These observations suggest that calcineurin and protein kinase C perform independent but physiologically related functions in yeast cells. We show that several mutants that lack a functional vacuolar H(+)-ATPase (vma) require calcineurin for vegetative growth. We discuss possible roles for calcineurin in regulating intracellular ion homeostasis and in maintaining cell integrity.
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PMID:Calcineurin, the Ca2+/calmodulin-dependent protein phosphatase, is essential in yeast mutants with cell integrity defects and in mutants that lack a functional vacuolar H(+)-ATPase. 754 41

Protein phosphatases regulate the activity of signal transduction mechanisms by dephosphorylating activated components. By utilizing selective inhibitors of these phosphatases, we investigated their role in regulating cAMP accumulation in the UMR 106 osteoblast-like tumor cell line. PTHrP, PTH and PGE2 stimulated cAMP accumulation up to 100-fold. Calyculin A, a potent inhibitor of protein phosphatase type 1 (PP1) and type 2A (PP2A), did not affect basal levels of cAMP, but concentrations of 10(-11) M to 10(-8) M increased PTHrP-, PTH-, and PGE2-stimulated cAMP accumulation up to 1.7-fold, and this increase was concentration-dependent. Similar results were obtained with tautomycin, another potent inhibitor of PP1 and PP2A. In contrast, okadaic acid, a potent inhibitor of PP2A which inhibited PP1 less potently, did not enhance PTHrP-, PTH-, or PGE2-stimulated cAMP accumulation. The effect of calyculin A on agonist-stimulated cAMP accumulation persisted in cells treated with isobutyl methylxanthine, a phosphodiesterase inhibitor. When the effect of calyculin A was compared with that of 4 beta-phorbol 12-myristate 13-acetate (PMA), it was found that while PMA enhanced both the receptor and forskolin-stimulated cAMP accumulation, calyculin A had no effect on the forskolin-stimulated cAMP accumulation. The effect of calyculin A on PTHrP- and PTH-stimulated cAMP accumulation persisted in cells treated with PMA. These results suggest that protein phosphatases play an important role in agonist-stimulated cAMP accumulation in osteoblast-like cells, and that PP1 but not PP2A may be the major phosphatase involved. In contrast to activation by protein kinase C, the site of action for the phosphatase appears to be predominantly at a step prior to the activation of adenylyl cyclase in the cAMP signal transduction pathway.
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PMID:Inhibition of serine/threonine protein phosphatases enhances agonist-stimulated cAMP accumulation in UMR 106 osteoblast-like cells. 754 25

The GTP-binding protein, G(o), is present at very high concentration in the neuronal growth cone membrane. The expression of activated mutants of the a subunit of G(o) increases neurite outgrowth. To determine the intracellular mechanism for this outgrowth, we have examined activated alpha o-dependent outgrowth in the presence of agents which modulate different signal transduction cascades. Activation of protein kinase C with phorbol esters or with diacylglycerol prevents the alpha o-dependent increase in neurite extension. Inhibition of protein kinase C with staurosporine, with H7, or with long-term, high dose phorbol ester treatment resulted in greater neurite elongation, and no further increase after activated alpha o transfection. The protein phosphatase inhibitor, okadaic acid, also blocked the effect of activated alpha o. In contrast, tyrosine kinase inhibitors and agents which alter cAMP levels did not alter activated alpha o-dependent neurite extension. We tested a number of compounds which alter intracellular calcium levels. TMB-8 and thapsigargin prevented an increase in outgrowth by activated alpha o, but diltiazem, Bay K8644 and dantrolene had no effect on activated alpha o-dependent outgrowth. These studies suggest that activated alpha o increases neurite outgrowth by inhibiting protein kinase C and by modulating intracellular calcium release.
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PMID:An activated mutant of the alpha subunit of G(o) increases neurite outgrowth via protein kinase C. 755 35

The MPK1 (SLT2) gene of Saccharomyces cerevisiae encodes a mitogen-activated protein kinase that is regulated by a kinase cascade whose known elements are Pkc1 (a homolog of protein kinase C), Bck1 (Slk1) (a homolog of MEK kinase), and the functionally redundant Mpk1 activators Mkk1 and Mkk2 (homologs of MEK). An activated mutation of MKK1, MKK1P386, inhibits growth when overexpressed. This growth-inhibitory effect was suppressed by the mpk1 delta mutation, suggesting that hyperactivation of the Mpk1 pathway is toxic to cells. To search for genes that interact with the Mpk1 pathway, we isolated both chromosomal mutations and dosage suppressor genes that ameliorate the growth-inhibitory effect of overexpressed Mkk1P386. One of the genes identified by the analysis of chromosomal mutations is RLM1 (resistance to lethality of MKK1P386 overexpression), which encodes a protein homologous to a conserved domain of the MADS (Mcm1, Agamous, Deficiens, and serum response factor) box family of transcription factors. Although rlm1 delta cells grow normally at any temperature, they display a caffeine-sensitive phenotype similar to that observed in mutants defective in BCK1, MKK1/MKK2, or MPK1. A gene fusion that provides Rlm1 with a transcriptional activation domain of Gal4 suppresses bck1 delta and mpk1 delta. A screening for dosage suppressors yielded the MSG5 genes, which encode a dual-specificity protein phosphatase. Our results suggest that Rlm1 functions as a transcription factor downstream of Mpk1 that is subject to activation by the Mpk1 mitogen-activated protein kinase pathway.
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PMID:Yeast RLM1 encodes a serum response factor-like protein that may function downstream of the Mpk1 (Slt2) mitogen-activated protein kinase pathway. 756 26

1. Activation of human D2(s) dopamine receptors with quinpirole (10 nM) inhibits omega-conotoxin GVIa-sensitive, high-threshold calcium currents when expressed in differentiated NG108-15 cells (55% inhibition at +10 mV). This inhibition was made irreversible following intracellular dialysis with the non-hydrolysable guanosine triphosphate analogue GTP-gamma-S (100 microM), and was prevented by pretreatment with pertussis toxin (1 microgram ml-1 for 24 h). 2. Stimulation of protein kinase C with the diacylglycerol analogue, 1-oleoyl-2-acetyl-sn-glycerol (100 microM), also attenuated the inhibition of the sustained calcium current but did not affect the receptor-mediated decrease in rate of current activation. Similarly, okadaic acid (100 nM), a protein phosphatase 1/2A inhibitor, selectively occluded the inhibition of the sustained current. 3. The depression of calcium currents by quinpirole (10 nM) was enhanced following intracellular dialysis with 100 microM cyclic adenosine monophosphate (cyclic AMP, 72.8 +/- 9.8% depression), but was not mimicked by the membrane permeant cyclic GMP analogue, Sp-8-bromoguanosine-3',5':cyclic monophosphorothioate (100 microM). 4. Inhibition of calcium currents was only partly attenuated by 100 ms depolarizing prepulses to +100 mV immediately preceding the test pulse. However, following occlusion of the sustained depression with okadaic acid (100 nM) the residual kinetic slowing was reversed in a voltage-dependent manner (P < 0.05). 5. Thus pertussis toxin-sensitive G-proteins liberated upon activation of human D2(short) dopamine receptors inhibited high-threshold calcium currents in two distinct ways. The decrease in rate of calcium current activation involved a voltage-dependent pathway, whereas the sustained inhibition of calcium current involved, in part, the voltage-resistant phosphorylation by cyclic AMP-dependent protein kinases and subsequent dephosphorylation by protein phosphatases 1/2A.
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PMID:Phosphorylation- and voltage-dependent inhibition of neuronal calcium currents by activation of human D2(short) dopamine receptors. 758 57

1. The mouse AtT-20/D16-16 anterior pituitary tumour cell line was used as a model system for the study of protein phosphatase involvement in the late stages of the secretory pathway for adrenocorticotrophin (ACTH) secretion. The effects of the type 1 and 2 phosphatase inhibitor calyculin A upon calcium-, guanine nucleotide- and phorbol 12-myristate 13-acetate (PMA)-stimulated ACTH secretion from electrically-permeabilized AtT-20 cells were studied. 2. Calyculin A (1 nM-1 microM) inhibited both calcium (10 microM)- and guanosine 5'-O-(3-thiotriphosphate) (GTP-gamma-S) (100 microM)-evoked ACTH secretion from permeabilized cells in a concentration-dependent manner. These effects were maximal with 100 nM calyculin A. 3. ACTH secretion was stimulated from electrically-permeabilized cells when the cytosolic free calcium ion concentration, controlled by calcium-EGTA buffers, was raised over the concentration range of 100 nM to 10 microM. This calcium-stimulated ACTH secretion was inhibited by co-incubation with calyculin A (100 nM). 4. GTP-gamma-S (10 nM-100 microM) stimulated ACTH secretion from permeabilized cells at concentrations greater than 1 microM GTP-gamma-S. Co-incubation with calyculin A (100 nM) inhibited this stimulation of ACTH secretion observed at these concentrations of GTP-gamma-S. 5. PMA (100 nM) significantly stimulated ACTH secretion from permeabilized cells in the absence of either calcium and guanine nucleotides and this action was enhanced by calyculin A (100 nM). Furthermore, an inhibition of GTP-gamma-S (100 microM)-stimulated ACTH secretion observed in the presence of calyculin A (100 nM) was not observed in the presence of PMA (100 nM). 6. The results of the present study indicate that dephosphorylation by phosphatases plays an important role in stimulus-secretion coupling in AtT-20 cells and is involved in mediating the effects of GE upon the secretory apparatus in these cells. Furthermore, the point of regulation of the secretory response by PKC which underlies the ability of PKC to amplify the calcium/GE system may lie distal to both GE and these phosphatases.
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PMID:The effects of calyculin A upon calcium-, guanine nucleotides- and phorbol 12-myristate 13-acetate-stimulated ACTH secretion from AtT-20 cells. 759 24


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