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

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Query: EC:2.7.11.1 (protein kinase)
81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have recently shown the selective inhibition of an amiloride-sensitive, conductive pathway for Na+ by atrial natriuretic peptide and 8-bromoguanosine 3',5'-cyclic monophosphate (8-BrcGMP) in the renal epithelial cell line, LLC-PK1. Using 22Na+ fluxes, we further investigated the modulation of Na+ transport by atrial natriuretic peptide and by agents that increase cGMP production, activate protein kinase c, or modulate guanine nucleotide regulatory protein function. Sodium nitroprusside increases intracellular cGMP concentrations without affecting cAMP concentrations and completely inhibits amiloride-sensitive Na+ uptake in a time- and concentration-dependent manner. In contrast, 8-BrcAMP is without effect on Na+ uptake through the Na+ channel. 1-Oleoyl 2-acetylglycerol (10 micrograms/ml) and phorbol 12-myristate 13-acetate (100 nM), activators of protein kinase c, inhibit Na+ uptake by 93 +/- 13 and 51 +/- 10%, respectively. Prolonged incubation with phorbol ester results in the downregulation of protein kinase c activity and reduces the inhibitory effect of atrial natriuretic peptide, suggesting that the action of this peptide involves stimulation of protein kinase c. Pertussis toxin, which induces the ADP-ribosylation of a 41-kDa guanine nucleotide regulatory protein in LLC-PK1 cells, inhibits 22Na+ influx to the same extent as amiloride. Thus, increasing cGMP, activating protein kinase c, and ADP-ribosylating a guanine nucleotide regulatory protein all inhibit Na+ uptake. These events may be sequentially involved in the action of atrial natriuretic peptide.
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PMID:Inhibition of epithelial Na+ transport by atriopeptin, protein kinase c, and pertussis toxin. 295 93

Using purified rat ventricular myocytes and membranes prepared from them, we have previously found that alpha 1-adrenergic stimulation causes decreased cyclic AMP accumulation and decreased activation of cyclic AMP-dependent protein kinase. We have now analyzed the mechanism by which alpha 1 stimulation is linked to cyclic AMP metabolism. In an adenylate cyclase assay in which carbachol inhibits the stimulatory effect of norepinephrine, the addition of prazosin (alpha 1-antagonist) has no effect on the response to norepinephrine. In membranes prepared from myocytes treated with pertussis toxin, norepinephrine competes for alpha 1-receptors (assessed by [3H]prazosin binding) with two components, binding to the high affinity component being sensitive to exogenous GTP, exactly as in membranes prepared from control myocytes. In intact cells labeled with [3H]adenine in which carbachol antagonizes the norepinephrine response, prazosin enhances accumulation of [3H]cyclic AMP due to norepinephrine. Treatment of cells with pertussis toxin eliminates inhibition by carbachol but does not alter prazosin's capacity to enhance the norepinephrine response. Addition of phosphodiesterase inhibitors eliminates this effect of alpha 1 blockade. In [3H]adenine-labeled cells loaded with [3H]cyclic AMP by prior treatment with isoproterenol, alpha 1-adrenergic stimulation enhances disappearance of [3H]cyclic AMP. Measurements of cellular cyclic AMP give results similar to those obtained with the adenine labeling technic. We conclude that occupation of the myocyte alpha 1-receptor results in stimulation of cyclic AMP phosphodiesterase activity.
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PMID:Action of the cardiac alpha 1-adrenergic receptor. Activation of cyclic AMP degradation. 298 30

Sarcolemmal membranes were isolated from porcine skeletal muscle by modifications of a LiBr-extraction technique. Latency determinations of acetylcholinesterase, ouabain-sensitive p-nitrophenylphosphatase, [3H]ouabain binding, and (Na+ + K+)-ATPase activities indicated that 65-76% of the membranes were sealed inside-out vesicles. The preparations were enriched in cholesterol and phospholipid, and demonstrated adenylate cyclase activity and both cAMP and cGMP phosphodiesterase activities. An indication of the purity of this fraction was that the Ca2+-ATPase activity (0.13 mumol Pi mg-1 min-1 at 37 degrees C) was 3.8% of that of porcine skeletal muscle sarcoplasmic reticulum preparations. Pertussis toxin specifically catalyzed the ADP-ribosylation of a Mr 41,000 sarcolemmal protein, indicating the presence of the inhibitory guanine nucleotide regulatory protein of adenylate cyclase, Ni. An endogenous ADP-ribosyltransferase activity, with several membrane protein substrates, was also demonstrated. The addition of exogenous cAMP-dependent protein kinase or calmodulin promoted the phosphorylation of a number of sarcolemmal proteins. The calmodulin-dependent phosphorylation exhibited an approximate K 1/2 for Ca2+ of 0.5 microM, and an approximate K 1/2 for calmodulin of 0.1 microM. 125I-Calmodulin affinity labeling of the sarcolemma, using dithiobis(succinimidyl propionate), demonstrated the presence of Mr 160,000 and 280,000 calmodulin-binding components in these membranes. These results demonstrate that this porcine preparation will be valuable in the study of skeletal muscle sarcolemmal ion transport, protein and hormonal receptors, and protein kinase-catalyzed phosphorylation.
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PMID:Components of purified sarcolemma from porcine skeletal muscle. 299 26

Adipocytes from hypothyroid rats have a decreased responsiveness to agents that activate adenylate cyclase, whereas cells from hyperthyroid rats have an increased responsiveness as compared to the controls. This is reflected in cyclic AMP accumulation as well as lipolysis. Administration of pertussis toxin to rats or its in vitro addition to adipocytes increased basal lipolysis and cyclic AMP accumulation as well as the response to norepinephrine or forskolin. The effects of thyroid status was not abolished by toxin treatment. Pertussis toxin-catalyzed ADP ribosylation of Ni was increased in adipocyte membranes from hypothyroid rats as compared to those from euthyroid rats. However, no change in sensitivity to N6-(phenylisopropyl)adenosine was observed. The data suggest that the amount of Ni might not be rate-limiting for the inhibitory action of adenosine. A consistent decrease in maximal lipolysis was observed in freshly isolated adipocytes from hypothyroid animals as compared to those from the controls. Such defective maximal lipolysis was not corrected by adenosine deaminase or in vivo administration of pertussis toxin. The relationship between cyclic AMP levels and lipolysis suggests that in fat cells from hypothyroid rats either the cyclic AMP-dependent protein kinase or the lipase activity itself may limit maximal lipolysis. There appears to be multiple effects of thyroid status on lipolysis involving factors other than those affecting adenylate cyclase activation.
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PMID:Pertussis toxin effects on adenylate cyclase activity, cyclic AMP accumulation and lipolysis in adipocytes from hypothyroid, euthyroid and hyperthyroid rats. 301 Nov 6

Intact platelets were stimulated with thrombin and the amount of GTP-binding protein (G-protein) oligomers was assessed by measuring ADP ribosylation of 40-41 kDa protein by pertussis toxin in isolated membranes. The toxin substrate fell by 57-62% in 10-60 s, but then returned towards normal over 5 min. Recovery was greatly enhanced by removal of thrombin from receptors with hirudin. Phorbol myristate acetate increased ADP-ribosylatable protein, but only back to initial levels prior to PMA. In contrast prostaglandin D2 plus theophylline (which increase cyclic AMP) did not increase ADP ribosylation, but could completely block the fall of the toxin substrate caused by thrombin. These results indicate that activation of thrombin receptors promotes the dissociation of G-protein oligomers to release free alpha-subunits, and this effect can be modulated by protein kinase C and cyclic AMP-dependent protein kinase. The possible relationships of these findings to the regulation of stimulus-response coupling in platelets is discussed.
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PMID:Effects of thrombin, phorbol myristate acetate and prostaglandin D2 on 40-41 kDa protein that is ADP ribosylated by pertussis toxin in platelets. 301 84

Pertussis toxin (PT), which blocks the activity of several G-proteins, has been found to exert a marked inhibitory effect on the DNA synthesis induced in 3T3 cells by serum or growth factors. 3T3 cells transformed with human c-ras oncogenes (Ha-ras, Ki-ras, N-ras) or with src, an oncogene coding for a protein kinase, have lost sensitivity to growth control by PT, even though substrates for PT can still be ADP-ribosylated in vivo. In contrast, 3T3 cells transformed with the SV40 virus behave like normal untransformed cells with respect to the ability of PT to decrease their growth rate. Oncogenes can thus likely be classified either as 'responders' or 'non-responders' to PT.
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PMID:Differential sensitivity to pertussis toxin of 3T3 cells transformed with different oncogenes. 304 49

1. The receptor-activated mechanisms that mediate the steroidogenic actions of angiotensin II (AII) have been characterized in rat and bovine adrenal glomerulosa cells. In rat adrenal cells, the AII receptor is coupled to a guanine nucleotide inhibitory protein which reduces adenylate cyclase activity and cyclic AMP production. However, receptor-mediated stimulation of aldosterone production by AII is exerted through a separate pertussis-insensitive nucleotide regulatory protein that subserves coupling of activated receptors to phospholipase C. 2. In AII-stimulated glomerulosa cells, hydrolysis of phosphatidylinositol (4,5)-bisphosphate (PIP2) by phospholipase C yields diacylglycerol and inositol 1,4,5-trisphosphate (Ins-P3), which act as second messengers by activating calcium-calmodulin and calcium-phospholipid dependent protein kinase pathways. Ins-1,4,5-P3 is a potent stimulus of intracellular calcium mobilization, and is promptly inactivated by two major routes of metabolism. Direct degradation of Ins-1,4,5-P3 by a 5-phosphatase gives inositol 1,4-bisphosphate which in turn is metabolized to inositol-4-monophosphate. The latter product can be derived only from higher inositol phosphates, and thus serves as a specific marker of polyphosphoinositide breakdown in agonist-stimulated cells. In contrast, inositol-1-phosphate is largely derived from phosphatidylinositol hydrolysis, which is not increased during the initial phase of AII action. 3. Ins-1,4,5-P3 formed in AII-stimulated glomerulosa cells is also phosphorylated by a calcium-calmodulin dependent 3-kinase to form inositol 1,3,4,5-tetrakisphosphate (Ins-P4), which is rapidly dephosphorylated to the biologically inactive Ins-1,4,5-P3 isomer, Ins-1,3,4-trisphosphate. The latter metabolite, like Ins-1,4,5-P3, is both degraded to lower phosphates (Ins-3,4,P2 and Ins-1,3-P2) and phosphorylated to form a new tetrakisphosphate isomer (Ins-1,3,4,6-P4). Ins-1,4,5-P3 formed during AII action is bound with high affinity to specific intracellular receptors through which InsP3 causes calcium mobilization during the initiation of cellular responses to AII and other calcium-dependent ligands.
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PMID:Control of glomerulosa cell function by angiotensin II: transduction by G-proteins and inositol polyphosphates. 315 62

Recent advances in insulin secretion indicate that pertussis toxin abolishes the inhibition by alpha 2 adrenoceptor activation of insulin release by the pancreas. Pertussis toxin adenosine diphosphate (ADP) ribosylates an inhibitory guanine nucleotide-binding protein (Ni) involved in inhibition of adenylate cyclase. The decrease in cyclic adenosine monophosphate (AMP) by epinephrine may account for its inhibition of insulin release. Insulin interaction with its receptor results in an increase in the tyrosine protein kinase activity of the receptor. Second messengers for insulin are generated, hexose transport is accelerated, and a cyclic AMP-independent protein kinase is activated that phosphorylates at serinethreonine residues. The activity of membrane-bound enzymes such as adenylate cyclase and Ca2+-Mg2+-ATPase is affected. The relative importance of these effects of insulin in its regulation of cellular metabolism remains to be established.
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PMID:Insulin secretion and action. 614 90

1. Whole-cell patch-clamp technique was used to study the beta-adrenergic and cholinergic regulation of the inwardly rectifying K+ conductance (gK1) in isolated guinea-pig ventricular myocytes. 2. In Cl(-)-free solutions or in the presence of 9-anthracenecarboxylic acid or Co2+, bath-applied isoprenaline (Iso) partially inhibited the steady-state whole-cell conductance (gss) calculated from the steady-state current (Iss)-voltage (Iss-V) curve at membrane voltages (Vm) negative to the equilibrium potential for potassium (EK). Iss was also inhibited at Vm positive to EK when the extracellular [K+] was 20 mM. The Iso-sensitive component of gss exhibited the characteristics of the inwardly rectifying K+ conductance (gK1). 3. The Iso-induced inhibition of gK1 was reversible, concentration dependent, blocked by propranolol, mimicked by both forskolin and dibutyryl cAMP, and prevented by including a cAMP-dependent protein kinase (PKA) inhibitor in the pipette solution. These findings suggest that PKA mediates the Iso-induced inhibition of gK1. 4. The apparent dissociation constant (KD) for the concentration dependence of Iso-induced inhibition was 0.035 microM and the Hill coefficient was approximately 1.0. A maximal Iso concentration (1 microM) inhibited gK1 by 40 +/- 4.1% (mean +/- S.E.M.; n = 13). 5. Bath application of acetylcholine (ACh, 0.1 microM or more) antagonized the Iso-induced (1 microM) inhibition of gK1; [ACh] > 1.0 microM antagonized 88 +/- 2.1% (n = 10) of the inhibition. ACh increased the KD for Iso to inhibit Iso-sensitive gK1 and also reduced the maximal Iso-induced inhibition. 6. ACh-induced antagonism could be abolished by pre-incubating myocytes with pertussis toxin (PTX), suggesting that a muscarinic receptor-coupled, PTX-sensitive G protein, Gi, is involved. 7. ACh (10 microM) also antagonized approximately 70% of the dibutyryl cyclic AMP (1 mM)-induced inhibition of gK1 (n = 3), suggesting that the ACh-induced antagonism involves more than simply inhibiting the Iso-mediated activation of adenylyl cyclase via the activated Gi. 8. Intracellularly applied okadaic acid (OkA, 1 microM) did not alter gK1 (control = 134 +/- 5.1 nS vs. OkA = 136 +/- 6.1 nS), but the Iso-induced decrease in gK1 was less (P < 0.001) with OkA present (42.1 +/- 2.4 nS, n = 5) than when absent (54.0 +/- 2.2 nS, n = 10). However, ACh (10 microM) failed to antagonize Iso-induced inhibition with OkA present, suggesting involvement of a protein phosphatase.
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PMID:beta-adrenergic and cholinergic modulation of the inwardly rectifying K+ current in guinea-pig ventricular myocytes. 747 26

1. To clarify the nature of the inhibition of whole-cell inwardly rectifying K+ current (IK1) by isoprenaline (Iso) and its antagonism by acetylcholine (ACh), we studied the effects of Iso and ACh and their surrogates on single channel currents (iK1) carried by inwardly rectifying K+ channels in cell-attached and excised inside-out patches obtained from guinea-pig ventricular myocytes. 2. Bath application of Iso suppressed iK1 channel activity in cell-attached patches. This was inhibited by propranolol. Bath-applied forskolin or dibutyryl cAMP mimicked the effect of bath-applied Iso. 3. Exposure of the cytosolic face of inside-out patches to purified catalytic subunit of the cAMP-dependent protein kinase (PKA) also suppressed iK1 channel activity, mimicking the effect of bath-applied Iso on iK1 recorded from cell-attached patches. 4. When applied directly to cell-attached patches via the patch pipette solution, ACh antagonized Iso-induced (1 microM applied via the bath) suppression of iK1 channels. In contrast, bath-applied ACh (10 microM) partially antagonized the effect of low concentrations of Iso (e.g. < 50 nM) on iK1 channels in cell-attached patches but had no detectable effect when 1 microM or more Iso was used. 5. In myocytes pretreated with pertussis toxin (PTX), ACh failed to antagonize Iso-induced suppression of iK1 channels. When inside-out patches were used, bath-applied preactivated exogenous inhibitory G protein subunit, G1 alpha, antagonized the suppression of iK1 channels induced by bath-applied catalytic subunit of PKA (PKA-CS), suggesting that a PTX-sensitive G1 alpha mediates ACh-induced antagonism of Iso-induced suppression of iK1. 6. Neither GTP gamma S nor G1 alpha antagonized the suppression of iK1 produced by bath-applied PKA-CS in inside-out patches when okadaic acid was present in the bath. In addition, bath application of alkaline phosphatase also reactivated iK1 channels suppressed by PKA-CS. 7. Findings in guinea-pig ventricular myocytes suggest that iK1 can be suppressed by a PKA-mediated phosphorylation of the iK1 channel occurring in response to Iso-induced beta-adrenergic receptor activation and that ACh can antagonize the suppression by mechanisms that involve both intracellular and membrane-delimited pathways. The membrane-delimited pathway appears to involve M2-cholinergic receptors, their associated G protein, G1, and a protein phosphatase, all located in the sarcolemma in close proximity to the involved iK1 channels.
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PMID:Beta-adrenergic and cholinergic modulation of inward rectifier K+ channel function and phosphorylation in guinea-pig ventricle. 747 27


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