Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.3.1.21 (CPT)
 4,580 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The enzymes cAMP-dependent protein kinase (PKA) and protein kinase C (PKC) regulate the activity of cardiac ion channel proteins. In this study the whole-cell arrangement of the patch clamp technique was used to examine the effect of NaI on PKA-stimulated Cl- and Ca2+ channels in isolated guinea pig ventricular myocytes. Cl- currents (ICl) activated either by the beta-adrenergic agonist isoproterenol or the membrane-soluble cAMP analogue, 8-chlorphenylthio (8-CPT) cAMP, were greatly reduced in amplitude after substitution of an external solution containing 140 mM NaCl with a solution containing 140 mM NaI. This reduction was accompanied by a shift of -7 mV in the reversal potential (Erev) for ICl and could be reversed upon return to the NaCl external solution. Inhibition of ICl by NaI occurred in a concentration-dependent manner and was more pronounced for inward ICl (IC50 = 19 mM at -60 mV) than for outward ICl (IC50 = 60 mM at +60 mV). In contrast to ICl activated by PKA, ICl activated by PKC was slightly augmented in the presence of NaI and the Erev was found to shift by -15 mV. Based on these data, the relative permeability of I- to Cl- (PI/PCl) for this channel was calculated to be 1.79. NaI produced no change in the amplitude of inward calcium currents (ICa) recorded under basal conditions, but strongly inhibited ICa augmented by isoproterenol and 8-CPT cAMP, and during dialysis of cells with the catalytic subunit of PKA (CS). The in vitro incorporation of [gamma-32P]ATP into histone IIA and Kemptide, measured in the presence of PKA and cAMP, was not significantly different in assay mixtures containing salts of Cl- and I-. However, the ability of isoproterenol to augment basal ICa in whole-cell experiments was attenuated when experiments were carried out entirely in NaI external solution. Thus, the reduction in ICl and ICa observed in this study may result from a direct effect of I- on the phosphorylation/dephosphorylation of cardiac ion channel proteins or associated regulatory proteins.
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PMID:Inhibition of heart calcium and chloride currents by sodium iodide. Specific attenuation in cAMP-dependent protein kinase-mediated regulation. 128 46

Hepatocytes respond to stimulation by glycogenolytic agonists acting via phosphoinositide (PI) breakdown through oscillations of the free cytosolic concentration of Ca2+ ([Ca2+]cyt.). Since the second-messenger repertoire of hepatocytes includes many other factors besides Ca2+, we investigated to what degree the regulation of [Ca2+]cyt. oscillations is integrated into these other signalling systems. [Ca2+]cyt. was recorded in single rat hepatocytes by using the Ca(2+)-indicator fura-2. Parallel stimulation with phenylephrine (an alpha 1-adrenergic agonist of PI breakdown) and glucagon resulted in a synergistic stimulation of [Ca2+]cyt. oscillations. Direct activation of the cyclic-AMP-dependent pathway with several stimuli (forskolin, 8-bromo cyclic AMP, 8-CPT cyclic AMP) mimicked the response to glucagon. In contrast, [Ca2+]cyt. oscillations induced by various combinations of these agonists could be antagonized by the glycogenic hormone insulin. As one of the options in the insulin-signalling network, we tested a diacylglycerol activator of protein kinase C, DiC8. It also acted as an inhibitor of [Ca2+]cyt. oscillations. We investigated how these observations could be reconciled with our previously introduced model of [Ca2+]cyt. oscillations in hepatocytes [Somogyi and Stucki (1991) J. Biol. Chem. 266, 11068-11077]. First of all, the effect of calmodulin inhibitors (calmidazolium and CGS 9343 B), acting at the core of our model on the feedback of Ca2+ on Ins(1,4,5)P3-induced Ca2+ release, was not altered by the new modulators. In addition, all agonists and antagonists could be used interchangeably in combination and introduced no significant change in the oscillatory pattern or spike shape. Since the response was solely limited to frequency modulation, over- or understimulation of the oscillatory system, there is no need to create a new oscillator or to introduce further reaction steps into the core of the model. We conclude that the regulation of [Ca2+]cyt. via the explored second-messenger pathways can be embedded into the oscillatory system as modulation of rate constants already present in this model.
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PMID:Modulation of cytosolic-[Ca2+] oscillations in hepatocytes results from cross-talk among second messengers. The synergism between the alpha 1-adrenergic response, glucagon and cyclic AMP, and their antagonism by insulin and diacylglycerol manifest themselves in the control of the cytosolic-[Ca2+] oscillations. 132 20

Acidification of the endosomal pathway is important for ligand and receptor sorting, toxin activation, and protein degradation by lysosomal acid hydrolases. Fluorescent probes and imaging methods were developed to measure pH to better than 0.2 U accuracy in individual endocytic vesicles in Swiss 3T3 fibroblasts. Endosomes were pulse labeled with transferrin (Tf), alpha 2-macroglobulin (alpha 2M), or dextran, each conjugated with tetramethylrhodamine and carboxyfluorescein (for pH 5-8) or dichlorocarboxyfluorescein (for pH 4-6); pH in individual labeled vesicles was measured by ratio imaging using a cooled CCD camera and novel image analysis software. Tf-labeled endosomes acidified to pH 6.2 +/- 0.1 with a t1/2 of 4 min at 37 degrees C, and remained small and near the cell periphery. Dextran- and alpha 2M-labeled endosomes acidified to pH 4.7 +/- 0.2, becoming larger and moving toward the nucleus over 30 min; approximately 15% of alpha 2M-labeled endosomes were strongly acidic (pH less than 5.5) at only 1 min after labeling. Replacement of external Cl by NO3 or isethionate strongly and reversibly inhibited acidification. Addition of ouabain (1 mM) at the time of labeling strongly enhanced acidification in the first 5 min; Tf-labeled endosomes acidified to pH 5.3 without a change in morphology. Activation of phospholipase C by vasopressin (50 nM) enhanced acidification of early endosomes; activation of protein kinase C by PMA (100 nM) enhanced acidification strongly, whereas elevation of intracellular Ca by A23187 (1 microM) had no effect on acidification. Activation of protein kinase A by CPT-cAMP (0.5 mM) or forskolin (50 microM) inhibited acidification. Lysosomal pH was not affected by ouabain or the protein kinase activators. These results establish a methodology for quantitative measurement of pH in individual endocytic vesicles, and demonstrate that acidification of endosomes labeled with Tf and alpha 2M (receptor-mediated endocytosis) and dextran (fluid-phase endocytosis) is sensitive to intracellular anion composition, Na/K pump inhibition, and multiple intracellular second messengers.
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PMID:Second messengers regulate endosomal acidification in Swiss 3T3 fibroblasts. 138 79

We examined the effect of carbachol, an acetylcholine analogue, on hydraulic conductivity (Lp) response to 10 microU/ml arginine vasopressin (AVP) in rabbit cortical collecting duct (CCD). In CCDs in which water flow had been established with AVP, subsequent addition of carbachol caused Lp (X10(-7) cm.atm-1.s-1) to fall from 251 +/- 32 to 146 +/- 19. Carbachol washout resulted in recovery of Lp to 217 +/- 38. In CCDs in which water flow had been established using 10(-4) M 8-chlorophenylthioadenosine 3',5'-cyclic monophosphate (8-CPT-cAMP), addition of carbachol had no effect. These posttreatment studies suggest that carbachol's effects on modulating established water flow occur at a "pre-cAMP" step. With carbachol added first, AVP-induced Lp was reduced from 233 +/- 24 (controls) to 105 +/- 19 (carbachol-pretreated). Pretreatment with 10(-6) M atropine, a muscarinic receptor antagonist, totally reversed the inhibitory effect of carbachol, consistent with a receptor-mediated effect of carbachol. Carbachol pretreatment also inhibited 8-CPT-cAMP-induced Lp, indicating that carbachol's effects also occur at a "post-cAMP" step. Pretreatment with 10(-7) M staurosporine, a protein kinase C (PKC) inhibitor, reversed inhibitory effect of carbachol on AVP-induced Lp (193 +/- 26), suggesting that carbachol's effects are mediated by PKC. Intracellular calcium concentration [( Ca2+]i) was measured in fura-2-loaded CCDs. Carbachol also increased [Ca2+]i from 229 +/- 120 to 389 +/- 160 nM.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Muscarinic receptor activation inhibits AVP-induced water flow in rabbit cortical collecting ducts. 164 93

The regulation of the plasma membrane Ca2+ pump by hormones via phosphorylation in intact cells has not been clearly established. We now present evidence that the Ca2+ pump is phosphorylated on both serine and threonine residues in unstimulated and stimulated cultured rat aortic endothelial cells. Among the stimuli tested, the protein kinase C activator phorbol 12-myristate 13-acetate (PMA) was most potent and increased the level of phosphorylation threefold, while the cAMP-dependent protein kinase activator 8-(4-chlorophenylthio)-cAMP (CPT-cAMP) stimulated the phosphorylation 1.6-fold. Two-dimensional tryptic phosphopeptide maps of the Ca2+ pump from unstimulated and CPT-cAMP-stimulated cells have identical patterns (five phosphopeptides) while PMA-stimulated cells have three additional phosphopeptides. Isoproterenol-, ATP-, angiotensin II-, and bradykinin-stimulated cells also have increased levels of Ca2+ pump phosphorylation. Stimuli-induced phosphorylation of the Ca2+ pump was rapid (5-10 min) and was concomitant with stimulated calcium efflux from the same cells. This is the first direct evidence that the plasma membrane Ca2+ pump in intact cells is regulated by various hormones or agonists via cAMP-dependent protein kinase or protein kinase C phosphorylation.
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PMID:Hormone-induced phosphorylation of the plasma membrane calcium pump in cultured aortic endothelial cells. 165 40

The possible involvement of protein kinase C and/or a lipoxygenase product in the mechanism by which adenosine inhibits release of [3H]acetylcholine evoked by electrical pulses from [3H]choline-labelled hippocampal slices was examined. For comparison, the muscarinic autoreceptors were examined using carbachol. The order of potency of adenosine analogues (CHA = R-PIA greater than NECA much greater than CGS 21680, CV 1808) indicates that the adenosine receptor responsible is of the A1 subtype. Adenosine (10 microM) and R-PIA (0.1 microM) were virtually equiactive as inhibitors and were antagonized to an equal extent by 8-CPT with a potency (IC50 approximately 25 nM) which is also compatible with A1-receptor mediation. The effects of carbachol and of R-PIA were not antagonized by the lipoxygenase inhibitor NDGA (10 or 50 microM). Stimulation of protein kinase C by the phorbol ester 4 beta-phorbol 12,13-dibutyrate caused a concentration-dependent increase in stimulation-evoked 3H overflow, but did not antagonize the presynaptic inhibitory effect of R-PIA or carbachol (0.01-1 microM). Staurosporine (0.1 microM), which inhibited the stimulating effect of phorbol dibutyrate, did not alter the effects of carbachol or R-PIA. The presynaptic effects of phorbol dibutyrate, R-PIA and adenosine were reduced by pretreatment with N-ethylmaleimide (100 microM for 10 min), which inactivates G-proteins. The evoked transmitter release was unaffected by nifedipine (1 microM) in the presence and in the absence of phorbol dibutyrate. These results indicate that adenosine, by acting at presynaptic A1-receptors, reduces transmitter release by a mechanism that involves neither an NDGA-sensitive lipoxygenase nor protein kinase C. The results also indicate that the enhancement of transmitter release by phorbol esters is due to protein kinase C activation and that a G-protein may be involved in the effect but a dihydropyridine-sensitive L-type Ca2+ channel probably is not.
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PMID:Adenosine A1-receptor-mediated inhibition of evoked acetylcholine release in the rat hippocampus does not depend on protein kinase C. 226 53

The enzymes adenosine 3',5'-monophosphate (cAMP)-dependent protein kinase (protein kinase A) and protein kinase C regulate the activity of a diverse group of cellular proteins including membrane ion channel proteins. When protein kinase A was stimulated in cardiac ventricular myocytes with the membrane-soluble cAMP analog 8-chlorphenylthio cAMP (8-CPT cAMP), the amplitude of the delayed-rectifier potassium current (IK) doubled when recorded at 32 degrees C but was not affected at 22 degrees C. In contrast, modulation of the calcium current (ICa) by 8-CPT cAMP was independent of temperature with similar increases in ICa occurring at both temperatures. Stimulation of protein kinase C by phorbol 12,13-dibutyrate also enhanced IK in a temperature-dependent manner but failed to increase ICa at either temperature. Thus, cardiac delayed-rectifier potassium but not calcium channels are regulated by two distinct protein kinases in a similar temperature-dependent fashion.
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PMID:Regulation of a heart potassium channel by protein kinase A and C. 284 75

1. Actions of protein kinase C activators, 1,2-oleoylacetylglycerol (OAG) and 12-O-tetradecanoylphorbol-13-acetate (TPA), on the glutamate-mediated neuromuscular transmission in the mealworm, Tenebrio molitor, were studied by the microelectrode current-clamp and voltage-clamp techniques. 2. The activators OAG and TPA stimulate the evoked and spontaneous transmitter releases from the presynaptic terminal, as evidenced by an increase in the quantum content estimated by the number of failures of extracellular excitatory postsynaptic potentials (EPSPs), and in the frequency of miniature EPSPs. 3. Both OAG and TPA act on the postsynaptic membrane to enhance responses to the transmitter L-glutamate. Protein kinase C activators increased the apparent maximum of the ionophoretic dose-response curve for glutamate-induced depolarization, without affecting the reversal potential and the voltage-dependent decay rate for the excitatory postsynaptic current (EPSC) under voltage-clamp conditions. 4. The postsynaptic effect of OAG and TPA is distinctly different from that of activators of cyclic nucleotide-dependent protein kinases, such as octopamine, forskolin, CPT-cyclic AMP (8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate), and 8-bromo-cyclic GMP (8-bromoguanosine 3',5'-cyclic monophosphate) which decreased the postsynaptic sensitivity to L-glutamate. 5. I suggest that the responsiveness of the receptor to L-glutamate is under the control of these counteracting enzyme systems in the insect neuromuscular junction.
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PMID:Activation of protein kinase C promotes glutamate-mediated transmission at the neuromuscular junction of the mealworm. 290 91

Incubation of rat hepatocytes with anandamide (arachidonoylethanolamide) inhibited acetyl-CoA carboxylase activity and fatty acid synthesis de novo without affecting fatty acid synthase. This was concomitant to a decrease in the intracellular levels of malonyl-CoA. Likewise, anandamide depressed both cholesterol synthesis de novo and the incorporation of exogenous palmitate into triacylglycerols and phospholipids. On the other hand, anandamide stimulated in parallel both carnitine palmitoyltransferase I activity and ketogenesis from palmitate, though ketogenesis from octanoate was unaffected. The effects of anandamide on hepatic fatty acid synthesis and oxidation were: (a) mimicked by arachidonic acid, a product of anandamide breakdown by anandamide amidase; (b) prevented by phenylmethylsulfonyl fluoride, an inhibitor of anandamide amidase; and (c) not affected by bisindolylmaleimide, a specific inhibitor of protein kinase C. Furthermore, delta 9-tetrahydrocannabinol had no effect on any of the parameters determined, ruling out the possibility that the effects of anandamide on hepatic fatty acid metabolism are mediated by the peripheral cannabinoid receptor. The results thus indicate that anandamide might function as a carrier of arachidonic acid in the modulation of hepatic fatty metabolism.
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PMID:Effects of anandamide on hepatic fatty acid metabolism. 757 52

LPS, a bacterial endotoxin, induces the expression of many genes in macrophages. We report the cloning of a novel 3.3-kb cDNA that is a member of the thymidylate kinase family of genes. This clone, which we have designated TYKi, was obtained by screening a cDNA library prepared from RNA isolated from the murine cell line RAW264.7 after bacterial LPS treatment. TYKi is quite similar to all thymidylate kinases for which there are sequence data. It conserves two very important domains in these kinases, namely, the catalytic domain or P-loop and the nucleotide binding domain. After LPS exposure, the TYKi message appears at 2 h, peaks at 6 h, and declines at 8 h. LPS induction of TYKi is dependent on de novo protein synthesis. Increasing cytosolic cAMP with forskolin attenuates the LPS induction of TYKi. However, treatment with 8-(4-chlorophenylthio)-cAMP (CPT-cAMP) or dibutyryl-cAMP did not affect the LPS induction of TYKi. In contrast, activation of protein kinase C with phorbol ester augmented the LPS response, whereas inhibiting protein kinase C with 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H7) suppressed the LPS response. Removing extracellular Ca2+ with EGTA inhibited LPS induction of TYKi, whereas increasing intracellular calcium with the calcium ionophore A23187 had little effect on the levels of the TYKi transcript. Inhibiting tyrosine kinase with genistein suppressed the induction of TYKi by LPS.
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PMID:A unique member of the thymidylate kinase family that is induced during macrophage activation. 775 51


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