EC:2.7.11.13 - FACTA Search

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)

The putative inhibitor of diacylglycerol kinase activity, 6-(2-[(4-fluorophenyl)phenylmethylene]-1-piperidinyl)-ethyl-7-meth yl-5H- thiazolo[3,2-a]pyrimidin-5-one (R59022), markedly potentiated cholecystokinin-C-terminal-octapeptide(CCK-8-)stimulated enzyme secretion from isolated rabbit pancreatic acini. Maximal potentiation occurred when acini were stimulated in the presence of 5-10 microM R59022. Potentiation depended both on the concentration of R59022 and CCK-8. No potentiation was observed when acini were half-maximally stimulated, whereas the secretory response to maximal and supramaximal concentrations of secretagogue was increased by 50-60%. R59022 alone had no effect on basal enzyme secretion and the drug did not potentiate the secretory response to the Ca2+ ionophore A23187 or to the phorbol ester, 12-O-tetradecanoylphorbol 13-acetate. Moreover, no increase in basal secretion was observed when acini were incubated in the presence of both R59022 and forskolin. These observations strongly suggest that receptor-mediated activation of the inositol phospholipid pathway is required for R59022-induced potentiation. R59022 inhibited the CCK-8-stimulated incorporation of 32Pi into phosphatidic acid dose dependently, without affecting the CCK-8-stimulated hydrolysis of 32P-labelled phosphatidylinositol 4,5-bisphosphate. This is consistent with an inhibitory effect of R59022 on acinar cell diacylglycerol kinase activity. The potentiating effect of R59022 was mimicked by 12-O-tetradecanoylphorbol 13-acetate added simultaneously with CCK-8. Therefore, it is concluded that in the presence of 5-10 microM R59022 the receptor-mediated increase in acinar cell diacylglycerol content is enhanced leading to enhanced activation of protein kinase C and to potentiation of the secretory response. The fact that the secretory response to maximal and supramaximal concentrations of CCK-8 is potentiated by R59022 suggests that at these concentrations of secretagogue the diacylglycerol/protein kinase C branch of the signal-transduction route is rate-limiting.
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PMID:The diacylglycerol kinase inhibitor, R59022, potentiates cholecystokinin-induced enzyme secretion from rabbit pancreatic acini. 169 Jun 50

Regulation of intracellular pH (pHi) was studied by dual wavelength fluorometry in monolayers of pancreatic AR42J cells loaded with the fluorescent probe 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein. In cells superfused with N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered solution at pH 7.40, basal pHi was determined to be 7.15 +/- 0.13. Na(+)-H+ exchange could be demonstrated in both resting cells and cells subjected to acid loading by use of transient exposure to NH4Cl. Na(+)-H+ exchange was completely blocked by 300 microM amiloride and was dependent on extracellular Na+ (apparent Km = 25 mM). When the concentration of the NH4Cl pulse was varied (0.5-25 mM), the rate of pHi recovery increased as pHi became acidic, reaching a maximum of 0.007 pH units/s at pHi of 6.4. Gastrointestinal hormones, including pentagastrin, cholecystokinin, and bombesin, increased the rate of Na(+)-H+ exchange without affecting cellular buffer capacity (21.5 +/- 1.8 mM/pH unit), thereby leading to an intracellular alkalinization. This was accompanied by a shift in the curve of Na(+)-H+ exchange as a function of pHi to more alkaline values, although the maximum rate of pH recovery was unchanged. Neither protein kinase C nor Ca2+ could be conclusively linked to activation of Na(+)-H+ exchange, raising the possibility of a more direct, receptor-controlled mechanism.
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PMID:Gastrointestinal peptides activate Na(+)-H+ exchanger in AR42J cells by increasing its affinity for intracellular H+. 169 2

The effects of two putative inhibitors of protein kinase C activity, staurosporine and H-7, on partially purified protein kinase C and amylase secretion from isolated rabbit pancreatic acini were investigated. Staurosporine dose-dependently inhibited amylase release stimulated by an optimal concentration of cholecystokinin C-terminal octapeptide. At a concentration of 100 nM, the drug inhibited the secretory response to the secretagogue by approximately 50%. At the same concentration, staurosporine inhibited 12-O-tetradecanoylphorbol 13-acetate-stimulated enzyme secretion by 90%. Moreover, the potentiating effect of this phorbol ester on cholecystokinin-induced amylase release was completely abolished in the presence of staurosporine. Interestingly, amylase release was decreased to the level observed with the combination of cholecystokinin and staurosporine. In contrast, H-7, potentiated rather than inhibited cholecystokinin-stimulated enzyme secretion, whereas the secretory response to 12-O-tetradecanoylphorbol 13-acetate was not affected by the drug. Both staurosporine and H-7, however, inhibited protein kinase C purified from exocrine pancreatic tissue. Kinetic analysis revealed that both compounds inhibited protein kinase C competitively with respect to ATP. The Ki value for staurosporine was 0.55 nM and for H-7 13.5 microM. Our results obtained with staurosporine are in line with a stimulatory role of protein kinase C in cholecystokinin-induced enzyme secretion from the exocrine pancreas. The results obtained with H-7 emphasize that care has to be taken in interpreting the biological effects of this drug.
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PMID:Dissimilar effects of the protein kinase C inhibitors, staurosporine and H-7, on cholecystokinin-induced enzyme secretion from rabbit pancreatic acini. 169 56

Recordings of [Ca2+]i in single AR42J cells loaded with Fura 2 were used to study regulation of [Ca2+]i oscillation. Continuous stimulation with the cholecystokinin analogue, (t-butyloxycarbonyl-Tyr-(SO3)-norleucine-Gly-Trp-Nle-Asp-2-phenylethyl ester) or carbachol evoked long lasting oscillation in [Ca2+]i. Removal of CCK-JMV-180 after brief stimulation did not abruptly stop the oscillation. Rather, removal of CCK-JMV-180 resulted in time-dependent reduction in amplitude with little change in frequency of oscillation. The patterns of [Ca2+]i oscillation were affected by activation of protein kinase C and protein kinase A. However, down-regulation of protein kinase C activity did not prevent stimulation of [Ca2+]i oscillation. Hence, we conclude that an active protein kinase C pathway is not crucial for [Ca2+]i oscillation in this cell line. Variation in extracellular Ca2+ concentration (Ca2+out) was used to further characterize the oscillation. Reducing Ca2+out to approximately 10 microM resulted in a time dependent inhibition of [Ca2+]i oscillation. Subsequent step increases in Ca2+out up to 2-3 mM resulted in increased amplitude and frequency of oscillation. Further increase in Ca2+out or an increase in plasma membrane permeability to Ca2+, brought about by an increase in pHo, resulted in increased amplitude, decreased frequency, and modified shape of the [Ca2+]i spikes. These observations point to the existence of regulatory mechanisms controlling the duration of Ca2+ release and entry during [Ca2+]i oscillation.
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PMID:Regulation of intracellular Ca2+ oscillation in AR42J cells. 170 Nov 71

Chornic exogenous administration of cholecystokinin octapeptide (CCK8) to rats led to a reduced sensitivity of pancreatic acinar cells to both CCK8 and carbachol stimulation without changes in affinity or number of CCK or muscarinic receptors. In addition, repeated feeding of camostate, a synthetic protease inhibitor which stimulates endogenous CCK release, desensitized the response of the acini to caerulein. This study investigates whether an altered postreceptor signal transduction mechanism is responsible for the reduced amylase secretion. Four days of camostate treatment significantly increased pancreatic weight, protein and amylase, but not DNA content, indicating organ hypertrophy, CCK8 and carbachol stimulated amylase release from acini, isolated from camostate-treated rats, was significantly reduced without shifting the dose response curve compared to controls. There was no difference in total phosphoinositide turnover between the groups. In addition, CCK8 and carbachol stimulated 45Ca efflux and calcium ionophore stimulated amylase release were similar in both groups. These results indicate that the release of calcium from intracellular stores and the utilization of intracellular calcium to drive amylase secretion is not affected in the hypertrophied pancreas. In contrast, incubation of acini from camostate-treated rats with TPA (a phorbol ester which directly stimulates protein kinase C) showed a 48% reduction in amylase secretion. This suggests that a regulatory mechanism is present at the level of protein kinase C or beyond, which is responsible for the decrease in amylase release in the hypertrophied pancreas.
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PMID:Intracellular mechanism responsible for reduced enzyme secretion from camostate-induced hypertrophied pancreas. 170 70

Treatment of rabbit pancreatic acini with the phorbol ester, 12-O-tetradecanoylphorbol 13-acetate (TPA), resulted in a time- and dose-dependent decrease of soluble protein kinase C activity coinciding with an increase of protein kinase C activity in the particulate fraction. After 5 min, soluble protein kinase C activity had decreased to almost 10% of the corresponding control. Total extractable protein kinase C activity, however, remained unchanged, indicating that the decrease of soluble protein kinase C activity was not due to TPA-induced inactivation of the enzyme. The biologically inactive phorbol ester, 4 alpha-phorbol 12,13-didecanoate, did not induce such a translocation of protein kinase C. The half-maximal concentration for TPA-induced translocation of protein kinase C was 40 nM, and was equal to that for TPA-induced amylase secretion from isolated acini. This suggests that translocation of protein kinase C to the particulate fraction is an important step in TPA-induced activation of protein kinase C and enzyme secretion. On the other hand, cholecystokinin, a secretagogue of the calcium-mobilizing type, whose secretory action is thought to be mediated, at least in part, by protein kinase C, did not change the subcellular distribution of protein kinase C. In the presence of R59022 6-(2-[(4-fluorophenyl)phenylmethylene]-1-piperidinyl ) ethyl-7-methyl-5H-thiazolo[3,2-a]pyrimidin-5-one, an inhibitor of diacylglycerol kinase activity, cholecystokinin produced a small but significant translocation of protein kinase C, suggesting that the inability of the hormone to induce translocation is not due to a rapid conversion of the diacylglycerol formed into phosphatidic acid.
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PMID:Effects of phorbol ester and cholecystokinin on the intracellular distribution of protein kinase C in rabbit pancreatic acini. 170 8

The mechanisms regulating the net synthesis of digestive enzymes during short-term stimulation by agonists were examined in pancreatic acini isolated from the rat. Dispersed pancreatic acini were stimulated for up to 60 min with various concentrations of cholecystokinin octapeptide (CCK-OP), carbachol, A23187, 4 beta-phorbol 12-myristate 13-acetate (PMA). The effects of these agonists on net protein synthesis was determined by measuring the incorporation of [3H]leucine or [35S]methionine into protein. Carbachol, PMA, A23187 and concentrations of CCK-OP of 100 pM and greater caused inhibition of protein synthesis. Fluorography of [35S]methionine labeled acinar cell proteins separated by one-dimensional SDS-polyacrylamide gel electrophoresis demonstrated that the agonists inhibited the synthesis of the digestive enzymes. Northern blot analysis using cDNA probes revealed that CCK-OP, carbachol and PMA did not alter the cellular content of amylase, lipase and elastase mRNA. The protein kinase C inhibitors 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H-7) and staurosporine failed to reverse the inhibitory effects of CCK-OP, carbachol and PMA on protein synthesis. CCK-OP and PMA activated phospholipase A (PLA) which liberated lysophosphatidylcholine (LPC) and free fatty acids from membrane phosphatidylcholine. Exogenously added PLA2 (Naja naja venom) inhibited protein synthesis and increased LPC to a similar extent as CCK and PMA. The results suggest that the inhibitory effects of CCK and carbachol on net protein synthesis are due to their effects on intracellular calcium and PLA-mediated breakdown of phosphatidylcholine rather than protein kinase C activation.
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PMID:Intracellular mechanisms involved in short-term regulation of net protein synthesis in pancreatic acini. 170 86

1. A comparative study was made of the effect of the phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA) on cholecystokinin octapeptide-evoked exocrine pancreatic secretion in the anaesthetized rat and isolated permeabilized pancreatic acinar cells. 2. Cholecystokinin octapeptide (CCK8; 0.10-6.40 nmol (kg body weight)-1) induced dose-dependent increases in pancreatic juice flow, total protein output and amylase release in the anaesthetized rat. 3. Administration of TPA (10(-8) mol (kg body weight)-1) in combination with CCK8 resulted in marked attenuation of the CCK8-evoked secretory response. 4. Simultaneous injection of polymyxin B (10(-8) mol (kg body weight)-1), an inhibitor of protein kinase C, with TPA and CCK8 reversed the inhibitory effect of the phorbol ester on CCK8-induced pancreatic juice flow, total protein output and amylase release. 5. In permeabilized rat pancreatic acini CCK8 (10(-13)-10(-9) M) elicited dose-dependent increases in [3H]leucine-labelled protein secretion (3H-labelled protein release). Combining TPA (10(-8) M) with CCK8 resulted in an inhibition of the CCK8-induced 3H-labelled protein release especially at lower concentrations of CCK8. At higher concentrations of CCK8, TPA was unable to inhibit the CCK8-evoked 3H-labelled protein release. Again, polymyxin B reversed the TPA-induced inhibition of CCK8-evoked 3H-labelled protein output. 6. The results indicate that protein kinase C activation may play an important physiological role in modulating the CCK8-evoked secretory response in rat pancreas in vivo and in vitro.
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PMID:Effects of phorbol ester on cholecystokinin octapeptide-evoked exocrine pancreatic secretion in the rat. 171 42

Regulation of cholecystokinin (CCK) and the proto-oncogene c-fos mRNA expression was studied in the human neuroblastoma cell line SK-N-MC. Cells were treated either with the tumor promoting phorbol-ester phorbol-12-myristate-13-acetate (PMA), the phosphodiesterase inhibitor isobutyl-methylxanthine (IBMX), which results in an elevated intracellular cyclic AMP (cAMP) level, or with a combination of PMA and IBMX. The level of CCK and c-fos mRNA was determined by Northern-blot analysis with CCK and c-fos specific antisense RNA probes after 4-24 h of drug treatment. Treatment with PMA and IBMX for 4-24 hours transiently raised the CCK mRNA level approximately 1.5-3.5 times compared to the controls, and the combination PMA and IBMX had an additive effect and elevated CCK mRNA abundance 1.5-6.5 times. Under the same experimental conditions, both PMA and IBMX elevated the c-fos mRNA level approximately 3-5.5 times. The drug combination showed a pronounced synergistic effect and raised the c-fos mRNA level approximately 3-20 times as compared to controls. Apparently, CCK and c-fos mRNA expression appears to be regulated by similar protein kinase C (PKC) and cAMP-dependent mechanisms in SK-N-MC cells.
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PMID:Phorbol 12-myristate-13-acetate (PMA) stimulates a differential expression of cholecystokinin (CCK) and c-fos mRNA in a human neuroblastoma cell line. 172 Apr 2

Cholecystokinin (CCK) and cholinergic agonists are both major stimulants of pancreatic enzyme secretion and both utilize a common calcium-phosphoinositide-mediated receptor coupling system. In this study we investigated the modulation of pancreatic acinar CCK receptors by the muscarinic agonist carbachol (CCh) and investigated the intracellular mechanisms involved in the modulation. Acini were isolated from rat pancreas and dispersed in N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-Ringer solution. Preincubation with 0.1 mM carbachol for 60 min reduced the CCK octapeptide (CCK-8; 100 pM)-stimulated amylase release by 43 +/- 5%. Binding of 125I-Bolton-Hunter-labeled CCK-8 (125I-BH-CCK-8) revealed two classes of CCK receptors, a high affinity with a dissociation constant (Kd) of 20 pM and a low affinity with a Kd of 2.3 nM. Pretreatment with 100 microM CCh decreased total binding by 35 +/- 6%, affecting the binding capacity of the high-affinity site, without change in the maximal binding capacity of the low-affinity site and no change in the Kd of either site. Preincubation of acini with 12-O-tetradecanoylphorbol 12,13-acetate (TPA, 1 microM), an activator of protein kinase C (PKC), decreased subsequent CCK-8-stimulated amylase release, and total binding of 125I-BH-CCK-8 to a similar extent as with pretreatment with CCh. The inhibitory effect of TPA or CCh on CCK-8-stimulated amylase release was reversed by simultaneous preincubation with H-7, an inhibitor of PKC. Pretreatment of acini with the calcium ionophore A23187, vasoactive intestinal peptide, or 8-bromoadenosine 3',5'-cyclic monophosphate had no effect on 125I-BH-CCK-8 binding. After CCh or TPA preincubation, CCK-8-stimulated production of [3H]inositol phosphates was inhibited by at least 49%.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Carbachol acts through protein kinase C to modulate cholecystokinin receptors on pancreatic acini. 172 47

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