Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.11.11 (AMPK)
 12,425 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The molecular basis of the regulation of cardiac L-type calcium channel activity by cAMP-dependent protein kinase (cA-PK) remains unclear. Direct cA-PK-dependent phosphorylation of the bovine ventricular alpha1 subunit in vitro has been demonstrated in microsomal membranes, detergent extracts and partially purified (+)-[3H]PN 200-110 receptor preparations. Two 32P-labeled phosphopeptides, derived from cyanogen bromide cleavage, of 4.7 and 9.5 kDa were immunoprecipitated specifically by site-directed antibodies against the rabbit cardiac alpha1 subunit amino acid sequences 1602-1616 and 1681-1694, respectively, consistent with phosphorylation at the cA-PK consensus sites at Ser(1627) and Ser(1700). No phosphopeptide products consistent with phosphorylation at three other C-terminal cA-PK consensus phosphorylation sites (Ser(1575), Ser(1848) and Ser(1928)) were identified using similar procedures suggesting that these sites are poor substrates for this kinase. Ser(1627) and Ser(1700) may represent sites of cA-PK phosphorylation involved in the physiological regulation of cardiac L-type calcium channel function.
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PMID:Cyclic AMP-dependent protein kinase phosphorylates residues in the C-terminal domain of the cardiac L-type calcium channel alpha1 subunit. 866 19

To explore the relationship between structure and function in the cystic fibrosis transmembrane conductance regulator (CFTR) Cl- channel, we studied Xenopus CFTR. We found that the anion permeability sequence of cAMP-activated Cl- currents in the apical membrane of Xenopus A6 epithelia differed from that of cAMP-activated Cl- currents in human epithelia expressing CFTR. To understand the molecular basis for this difference and to learn whether CFTR from another species would have properties similar to human CFTR, we assembled a full-length Xenopus CFTR cDNA from A6 cells. Expression of Xenopus CFTR in HeLa cells generated cAMP-activated whole-cell currents and cAMP-dependent protein kinase-activated single channels that resembled those of human CFTR with the exception that the anion permeability sequence was different (Br- = I- > Cl- in Xenopus CFTR and Br- = Cl- > I- in human). In addition, the single-channel conductance of Xenopus CFTR was increased. To investigate protein regions that account for these differences, we constructed chimeric proteins by replacing either the first or second membrane-spanning domain of human CFTR with the equivalent region of Xenopus CFTR (hX1-6 and hX7-12, respectively) and examined their function in HeLa cells. We found that the anion permeability sequence (Br- = I- > Cl-) and single-channel conductance of hX1-6 resembled that of Xenopus CFTR expressed in HeLa cells, whereas hX7-12 had properties like those of human CFTR. However, the gating of hX1-6 showed a flickery behavior. The altered gating of hX1-6 was attributed to residues in the first extracellular loop of Xenopus CFTR because mutation of residues in that region to the corresponding residues of human CFTR produced gating behavior similar to that of human CFTR. These data suggest that sequence differences in the first membrane-spanning domains are responsible for the differences in the permeation properties of human and Xenopus CFTR and that the first extracellular loop influences channel gating.
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PMID:Function of Xenopus cystic fibrosis transmembrane conductance regulator (CFTR) Cl channels and use of human-Xenopus chimeras to investigate the pore properties of CFTR. 881 Feb 76

(Rp)-8-Bromo-guanosine 3',5'-cyclic monophosphorothioate (Rp-8-Br-cGMPS) inhibited competitively both isozymes of type I alpha and I beta cGMP-dependent protein kinase (cGMP-kinase) purified from porcine aorta with apparent Ki values (microM) of 3.7 for I alpha and 1.8 for I beta. The compound also inhibited bovine heart type II cAMP-dependent protein kinase (cAMP-kinase), but with a Ki of 25 microM. Thus, it is a selective inhibitor of cGMP-kinase. In alpha-toxin-skinned smooth muscle preparations from rat mesenteric artery, 8-Br-cGMP (10(-7) M) and 8-Br-cAMP (10(-6) M) produced a rightward shift of the concentration-contraction curves for Ca2+, denoting a decrease in Ca2+ sensitivity of the contractile elements. The shift by 8-Br-cAMP as well as by 8-Br-cGMP was completely reversed by Rp-8-Br-cGMPS, while a selective inhibitor of activation of cAMP-kinase, (Rp)-adenosine-3',5'-cyclic monophosphorothioate (Rp-cAMPS), was without effects on the shift produced by these two compounds. These findings indicate the pivotal role that the activation of cGMP-kinase plays in the production of a decrease in Ca2+ sensitivity of contractile elements.
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PMID:cGMP-kinase mediates cGMP- and cAMP-induced Ca2+ desensitization of skinned rat artery. 910 79

The pericellular proteoglycan biglycan is among the major secretory products of osteoblasts and articular chondrocytes but the regulatory agents and signal transduction pathways that ultimately lead to alterations in biglycan gene expression are poorly defined. We report here on the transcriptional up-regulation of biglycan in MG-63 osteosarcoma cells by agents that increase intracellular cAMP levels. Transfection of these cells with biglycan promoter luciferase reporter fusion genes and subsequent treatment with forskolin or the cAMP analog 8-Bromo-cAMP resulted in an up to 3.8-fold stimulation of biglycan promoter activity. This effect could be prevented with the compound KT5720, a specific inhibitor of the cAMP-dependent protein kinase. Up-regulation of transcription is also reflected at the level of mRNA expression, since biglycan mRNA steady state levels in MG-63 cells increased approximately 2-fold after 24 hours of forskolin treatment. These data suggest that elevated levels of intracellular cAMP increase transcription from the biglycan promoter in bone cells and implicate for the first time the cAMP/protein kinase A signal transduction pathway in the regulation of biglycan gene expression.
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PMID:Biglycan gene promoter activity in osteosarcoma cells is regulated by cyclic AMP. 919 8

Purinoceptor agonists produced potassium currents with the order of potency: ATP > adenosine = ADP = AMP > beta,gamma-methylene ATP, while a small response or no response was induced by 2-methylthio ATP, UTP, or alpha,beta-methylene ATP. The response induced by beta,gamma-methylene ATP was completely inhibited in the presence of alpha,beta-methylene ATP, suggesting that the relevant receptor for these agonists was a P3 purinoceptor. ATP induced currents with a latency of 24 s and the currents were not induced in defolliculated oocytes. The currents were not affected by either the Gi/o-protein inhibitor, pertussis toxin (PTX), or the selective cAMP-dependent protein kinase inhibitor, H-89, or the phospholipase C (PLC) inhibitor, neomycin, or the phospholipase A2 (PLA2) inhibitor, 4-bromophenacyl bromide. The currents were enhanced by the selective protein kinase C (PKC) inhibitor, GF109203X, but otherwise, they were reduced by the potent PKC activator, 12-O-tetradecanoylphorbol-13-acetate. The results of the present study suggest that a P3 purinoceptor in the follicle cell layer of oocytes is involved in activation of potassium channels and that the evoked currents are regulated by PLC/PLA2-independent PKC activation.
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PMID:ATP produces potassium currents via P3 purinoceptor in the follicle cell layer of Xenopus oocytes. 965 60

To investigate the molecular mechanism(s) of action of catecholamines on the expression of the angiotensinogen (ANG) gene in kidney proximal tubular cells, we used opossum kidney (OK) cells with a fusion gene containing the 5'-flanking regulatory sequence of the rat ANG gene fused with a human growth hormone (hGH) gene as a reporter, pOGH (rANG N-1498/+18), permanently integrated into their genomes. The level of expression of the ANG-GH fusion gene was quantified by the amount of immunoreactive-hGH (IR-hGH) secreted into the medium. The addition of norepinephrine (NE), isoproterenol (a beta1/beta2-adrenergic receptor (AR) agonist) and iodoclonidine (an alpha2-AR agonist) stimulated the expression of the ANG-GH fusion gene in a dose-dependent manner, whereas the addition of epinephrine and phenylephrine (alpha1-AR agonist) had no effect. The stimulatory effect of NE was blocked by the presence of propranolol (beta-AR blocker), atenolol (beta1-AR blocker), yohimbine (alpha2-AR blocker), Rp-cAMP (an inhibitor of cAMP-dependent protein kinase AI & AII) and staurosporine (an inhibitor of protein kinase C), but was not blocked by ICI 118, 551 (beta2-AR blocker) and prazosin (alpha1-AR blocker). The addition of a combination of isoproterenol and iodoclonidine or a combination of 8-Bromo-cAMP (8-Br-cAMP) and phorbol 12-myristate (PMA) synergistically stimulated the expression of the ANG-GH fusion gene as compared to the addition of isoproterenol, iodoclonidine, 8-Br-cAMP or PMA alone. Furthermore, the addition of NE, 8-Br-cAMP or PMA stimulated the expression of pOGH (rANG N-806/-779/-53/+18), a fusion gene containing the putative cAMP responsive element (CRE, ANG N-806/-779) upstream of the ANG promoter (ANG N-53/+18) in OK cells, but had no effect on the expression of fusion genes containing the mutant of the CRE. Gel mobility shift assays revealed that the ANG-CRE binds with the DNA-binding domain (bZIP254-327) of the cAMP-responsive binding protein (CREB). The binding of the labeled ANG-CRE to CREB (bZIP254-327) was displaced by unlabeled ANG-CRE and the CRE of the somatostatin gene but not by the mutants of the ANG-CRE. Finally, NE stimulated the phosphorylation of CREB in OK cells. These studies demonstrate that the molecular mechanism(s) of NE action on the expression of the ANG gene in OK cells may be mediated via both the PKA and PKC signalling pathways and via the phosphorylation of CREB. The phosphorylated CREB then interacts with the CRE in the 5'-flanking region of the ANG gene and subsequently stimulates the gene expression.
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PMID:Catecholamines and angiotensinogen gene expression in kidney proximal tubular cells. 1110 38

1 Serotonin 5-HT(4) receptor isoforms differ in their C-terminal tail and yet little is known about their regulation. In this study, we investigated the desensitization of two human 5-HT(4) receptors stably expressed in CHO cells, with a special emphasis on the h5-HT(4(d)) isoform. 2 Exposure of h5-HT(4(d)) and h5-HT(4(e)) receptors to 1 micro M 5-HT induced a rapid desensitization of the adenylyl cyclase response. The h5-HT(4(d)) receptor desensitized with a faster rate (t(1/2)<5 min) than the h5-HT(4(e)) receptor (t(1/2)=15 min) and after 10 min 5-HT treatment cAMP production was reduced by approximately 70%. 3 5-HT-induced h5-HT(4(d)) receptor desensitization was mimicked by 8-Bromo-cAMP, a cAMP analogue, and was inhibited by [n-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulphonamide, 2HCl] (H-89), an inhibitor of cAMP-dependent protein kinase (PKA). Inhibitors of endocytosis (sucrose, 0.45 M and concanavaline A, 0.25 mg ml(-1)) partially reversed the h5-HT(4(d)) receptor desensitization process. 4 Given the prominent role of PKA in agonist-induced desensitization, we mutated the four putative PKA phosphorylation sites present in the third intracellular loop (Ser242, Thr253, Thr255) and the C terminal tail (Ser338) of the h5-HT(4(d)) receptor. Surprisingly, mutated receptors in which either one or all four putative phosphorylation sites were substituted to alanine did not impair receptor desensitization suggesting that PKA might act on nonconsensus sites. 5 Altogether, our data demonstrate that the C-terminal tail of h5-HT(4) receptors may influence the rate of agonist-induced desensitization and we provide evidence for a major role of PKA in h5-HT(4(d)) receptor desensitization.
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PMID:Characterization of human 5-HT4(d) receptor desensitization in CHO cells. 1256 69

This study tested the hypothesis that nitric oxide (NO) production contributes to relaxation induced by 3',5'-cyclic adenylate monophosphate (cAMP)-elevating agents and that high salt diet impairs this mechanism of relaxation. Relaxation response to isoproterenol but not sodium nitroprusside, a NO donor, was reduced in the thoracic aorta from rats that were placed on a high salt diet (8% NaCl; 60+/-4%, P<0.001). 1H-[1,2,4]oxadiazolol [4,3,-alpha]quinoxalin-1-one (ODQ, 10 microM), a soluble guanylate cyclase inhibitor, but not N(omega)-nitro-L-arginine methyl ester (L-NAME, 100 microM), an inhibitor of NO synthase (NOS), attenuated the relaxation to isoproterenol (59+/-16%, P<0.01). High salt diet also impaired the relaxation responses to forskolin, an activator of adenylate cyclase, or 8-Bromo-cAMP (8-Br-cAMP). (N-[2-((p-bromocinnamyl)aminoethyl]-5-isoquinolinesulfonamide hydrochloride (H-89) (8 microM), an inhibitor of cAMP-dependent protein kinase, did not affect the relaxation produced by isoproterenol. These data suggest that high salt diet impairs relaxation response to isoproterenol by a dual mechanism involving diminished NO/NOS pathway linked to cGMP pathway and diminished cAMP pathway that is independent of protein kinase A.
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PMID:High salt diet modulates cAMP- and nitric oxide-mediated relaxation responses to isoproterenol in the rat aorta. 1292 69

The host hormone melatonin increases cytoplasmic Ca(2+) concentration and synchronizes Plasmodium cell cycle (Hotta, C.T., M.L. Gazarini, F.H. Beraldo, F.P. Varotti, C. Lopes, R.P. Markus, T. Pozzan, and C.R. Garcia. 2000. Nat. Cell Biol. 2:466-468). Here we show that in Plasmodium falciparum melatonin induces an increase in cyclic AMP (cAMP) levels and cAMP-dependent protein kinase (PKA) activity (40 and 50%, respectively). When red blood cells infected with P. falciparum are treated with cAMP analogue adenosine 3',5'-cyclic monophosphate N6-benzoyl/PKA activator (6-Bz-cAMP) there is an alteration of the parasite cell cycle. This effect appears to depend on activation of PKA (abolished by the PKA inhibitors adenosine 3',5'-cyclic monophosphorothioate/8 Bromo Rp isomer, PKI [cell permeable peptide], and H89). An unexpected cross talk was found to exist between the cAMP and the Ca(2+)-dependent signaling pathways. The increases in cAMP by melatonin are inhibited by blocker of phospholipase C U73122, and addition of 6-Bz-cAMP increases cytosolic Ca(2+) concentration, through PKA activation. These findings suggest that in Plasmodium a highly complex interplay exists between the Ca(2+) and cAMP signaling pathways, but also that the control of the parasite cell cycle by melatonin requires the activation of both second messenger controlled pathways.
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PMID:Cyclic AMP and calcium interplay as second messengers in melatonin-dependent regulation of Plasmodium falciparum cell cycle. 1610 24

Using histamine and the H3 receptor antagonist thioperamide, the roles of histamine receptors in NMDA-induced necrosis were investigated in rat cultured cortical neurons. Within 3 h of intense NMDA insult, most neurons died by necrosis. Histamine reversed the neurotoxicity in a concentration-dependent manner and showed peak protection at a concentration of 10(-7) m. This protection was antagonized by the H2 receptor antagonists cimetidine and zolantidine but not by the H1 receptor antagonists pyrilamine and diphenhydramine. In addition, the selective H2 receptor agonist amthamine mimicked the protection by histamine. This action was prevented by cimetidine but not by pyrilamine. 8-Bromo-cAMP also mimicked the effect of histamine. In contrast, both the adenylyl cyclase inhibitor 9-(tetrahydro-2-furanyl)-9H-purine-6-amine and the cAMP-dependent protein kinase inhibitor N-[2-(p-bromocinnamylamino) ethyl]-5-isoquinolinesulfonamide reversed the protection by histamine. Thioperamide also attenuated NMDA-induced excitotoxicity, which was reversed by the H3 receptor agonist (R)-alpha-methylhistamine but not by pyrilamine and cimetidine. In addition, the protection by thioperamide was inhibited by the GABA(A) receptor antagonists picrotoxin and bicuculline. Further study demonstrated that the protection by thioperamide was due to increased GABA release in NMDA-stimulated samples. These results indicate that not only the H2 receptor/cAMP/cAMP-dependent protein kinase pathway but also the H3 receptor/GABA release pathway can attenuate NMDA-induced neurotoxicity.
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PMID:Histamine protects against NMDA-induced necrosis in cultured cortical neurons through H receptor/cyclic AMP/protein kinase A and H receptor/GABA release pathways. 1647 29


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