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)

Prenalterol, an allegedly beta 1-selective adrenergic agonist with high intrinsic sympathomimetic activity (ISA), was shown to be weakly lipolytic in rat adipocytes. However, in pertussis-toxin-treated adipocytes, the ISA of prenalterol was markedly increased (from 10-20% to approx. 100% of that of isoprenaline). The cellular sensitivity was also increased (EC50 approx. 60 nM and approx. 3 microM in pertussis-toxin-treated and control cells respectively). A similar effect was seen for other partial agonists such as the beta 2-selective agonist terbutaline and for beta-adrenergic antagonists with some intrinsic activity (metoprolol, pindolol). There was no clear change in sensitivity to isoprenaline's ability to stimulate adenylate cyclase in adipocyte membranes from pertussis-toxin-treated animals but the cyclase activity was increased approx. 4-fold in the presence of 1 microM-GTP. Prenalterol stimulated lipolysis by only small increases in intracellular cyclic AMP (cAMP) levels (less than 10% of that seen with isoprenaline). Basal lipolysis was increased in cells from pertussis-toxin-treated rats and the cellular sensitivity to the non-degradable cAMP analogue, N6-monobutyryl-cAMP, was increased. In control cells, a submaximal concentration of prenalterol (0.1 microM) increased the sensitivity to the cAMP analogues, N6-monobutyryl-cAMP and 8-bromo-cAMP. A low concentration (1 mM) of 8-bromo-cAMP also increased the effect of prenalterol. Similar effects were seen when the phosphodiesterase was inhibited. Thus (1) lipolysis is extremely sensitive to small increases in intracellular cAMP; (2) the degree of activation of adenylate cyclase and thus cAMP formation is the rate-limiting step for the biological response of partial agonists; (3) the inhibitory GTP-binding protein, Gi, is an important modulator ('tissue factor') of the beta-adrenergic agonistic property; (4) low levels of cAMP exert a priming effect on protein kinase A.
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PMID:The inhibitory GTP-binding protein (Gi) regulates the agonistic property of beta-adrenergic ligands in isolated rat adipocytes. Evidence for a priming effect of cyclic AMP. 128 Jan 15

Polyclonal (RP) and monoclonal (Ab) antibodies were raised against synthetic peptides (or fusion proteins) corresponding to amino acid sequences unique to human and mouse retinoic acid receptor-beta (RAR beta) isoforms. Antibodies directed against the A2 region [Ab6 beta 2(A2), Ab7 beta 2(A2), and RP beta 2(A2)], the D2 region [RP beta(D2)], or the F region [Ab8 beta(F)2, RP beta(F)1, and RP beta(F)2] were selected. The monoclonal and polyclonal antibodies directed against the D2 and F regions specifically immunoprecipitated and recognized by Western blotting all human and mouse RAR beta isoforms (mRAR beta 1, -beta 2, -beta 3, and -beta 4), produced in COS-1 cells transfected with expression vectors containing the corresponding RAR beta cDNA. Furthermore, in gel retardation assays, the monoclonal antibodies supershifted RAR beta protein-RA response element oligonucleotide complexes. Antibodies directed against the A2 region were specific for the RAR beta 2 isoform. The above antibodies allowed us to detect the presence of mRAR beta 2 proteins in mouse embryos and to show that their presence in embryonal carcinoma cells (F9 and P19 cell lines) is dependent upon RA treatment. The antibodies were also used to demonstrate that RAR beta proteins produced by transfection in COS-1 cells are phosphorylated. RAR beta 2 phosphorylation was not affected by RA treatment, whereas the phosphorylation of RAR beta 1 and RAR beta 3 isoforms was greatly enhanced by RA. We also show that, in contrast to RAR alpha 1 and RAR gamma 1, RAR beta 2 proteins contain phosphotyrosine residues and are only weakly phosphorylated in vitro by cAMP-dependent protein kinase. These results support our previous proposal that the various receptors have distinct functions in the RA-signaling pathway.
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PMID:Retinoic acid receptor-beta: immunodetection and phosphorylation on tyrosine residues. 128 41

We used the pH-sensitive fluorescent dye 2',7'-bis(2-carboxyethyl)-5(6')-carboxyfluorescein to monitor the recovery of the intracellular pH (pHi) of rat parotid acini from an NH4(+)-induced alkaline load. This recovery was markedly inhibited by the loop diuretic bumetanide and by Cl- removal, indicating that it is largely due to NH4+ entry via the basolateral Na(+)-K(+)-2Cl- cotransporter. The rate of recovery of pHi was enhanced threefold by pretreatment (37.5 s) with isoproterenol (K1/2 = 21.5 nM) or norepinephrine (in the presence of phentolamine), and blocked by the beta 1-specific antagonist atenolol, indicating an upregulation of cotransport activity by beta 1-adrenergic stimulation. The effect of isoproterenol was prevented by protein kinase inhibitors and mimicked by cAMP analogues, and by maneuvers known to increase cytosolic cAMP levels in these cells, consistent with the involvement of protein kinase A. Physiologically, such an upregulation of the acinar Na(+)-K(+)-2Cl- cotransporter would lead to an increase in acinar chloride uptake across the basolateral membrane, and consequently, an increase in overall chloride and fluid secretion. Prevention of this upregulation by beta-blockers and possibly by other commonly used clinical agents may account for the dry mouth and dry eyes experienced by some patients taking these medications.
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PMID:Beta-adrenergic upregulation of the Na(+)-K(+)-2Cl- cotransporter in rat parotid acinar cells. 131 47

Gamma-aminobutyric acid Type A (GABAA) receptors are the major sites of synaptic inhibition in the central nervous system. These receptors are thought to be pentameric complexes of homologous transmembrane glycoproteins. Molecular cloning has revealed a multiplicity of different GABAA receptor subunits divided into five classes, alpha, beta, gamma, delta, and rho, based on sequence homology. Within the proposed major intracellular domain of these subunits, there are numerous potential consensus sites for protein phosphorylation by a variety of protein kinases. We have used purified fusion proteins of the major intracellular domain of GABAA receptor subunits produced in Escherichia coli to examine the phosphorylation of these subunits by cAMP-dependent protein kinase (PKA) and protein kinase C (PKC). The purified fusion protein of the intracellular domain of the beta 1 subunit was an excellent substrate for both PKA and PKC. PKA and PKC phosphorylated the beta 1 subunit fusion protein on serine residues on a single tryptic phosphopeptide. Site-directed mutagenesis of serine 409 in the intracellular domain of the beta 1 subunit to an alanine residue eliminated the phosphorylation of the beta 1 subunit fusion protein by both protein kinases. The purified fusion proteins of the major intracellular domain of the gamma 2S and gamma 2L subunits of the GABAA receptor were rapidly and stoichiometrically phosphorylated by PKC but not by PKA. The phosphorylation of the gamma 2S subunit occurred on serine residues on a single tryptic phosphopeptide. Site-directed mutagenesis of serine 327 of the gamma 2S subunit fusion protein to an alanine residue eliminated the phosphorylation of the gamma 2S fusion protein by PKC. The gamma 2L subunit is an alternatively spliced form of the gamma 2S subunit that differs by the insertion of 8 amino acids (LLRMFSFK) within the major intracellular domain of the gamma 2S subunit. The PKC phosphorylation of the gamma 2L subunit occurred on serine residues on two tryptic phosphopeptides. Site-specific mutagenesis of serine 343 within the 8-amino acid insert to an alanine residue eliminated the PKC phosphorylation of the novel site in the gamma 2L subunit. No phosphorylation of a purified fusion protein of the major intracellular loop of the alpha 1 subunit was observed with either PKA or PKC. These results identify the specific amino acid residues within GABAA receptor subunits that are phosphorylated by PKA and PKC and suggest that protein phosphorylation of these sites may be important in regulating GABAA receptor function.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Identification of the cAMP-dependent protein kinase and protein kinase C phosphorylation sites within the major intracellular domains of the beta 1, gamma 2S, and gamma 2L subunits of the gamma-aminobutyric acid type A receptor. 132 Nov 50

gamma-Aminobutyric acidA (GABAA) receptors are ligand-gated ion channels that mediate inhibitory synaptic transmission in the central nervous system. The role of protein phosphorylation in the modulation of GABAA receptor function was examined with cells transiently transfected with GABAA receptor subunits. GABAA receptors consisting of the alpha 1 and beta 1 or the alpha 1, beta 1, and gamma 2 subunits were directly phosphorylated on the beta 1 subunit by adenosine 3',5'-monophosphate (cAMP)-dependent protein kinase (PKA). The phosphorylation decreased the amplitude of the GABA response of both receptor types and the extent of rapid desensitization of the GABAA receptor that consisted of the alpha 1 and beta 1 subunits. Site-specific mutagenesis of the serine residue phosphorylated by PKA completely eliminated the PKA phosphorylation and modulation of the GABAA receptor. In primary embryonic rat neuronal cell cultures, a similar regulation of GABAA receptors by PKA was observed. These results demonstrate that the GABAA receptor is directly modulated by protein phosphorylation and suggest that neurotransmitters or neuropeptides that regulate intracellular cAMP levels may modulate the responses of neurons to GABA and consequently have profound effects on synaptic excitability.
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PMID:Functional modulation of GABAA receptors by cAMP-dependent protein phosphorylation. 132 40

Calcium transport by the cardiac sarcoplasmic reticulum is depressed in human dilated cardiomyopathy, but the mechanisms involved are not clear. The possible involvement of immunological mechanisms was explored by evaluating the effect of sera from 49 patients with dilated cardiomyopathy on oxalate-facilitated Ca2+ uptake. In 14 of these patients, serum or IgG induced a time- and concentration-dependent decline (29 +/- 4% at 100-fold serum dilution) in Ca2+ transport. In 14 patients, autoantibodies against the beta 1-adrenoceptor were also demonstrated by a ligand binding inhibition assay. Serum from these patients inhibited the isoproterenol-mediated stimulation of Ca2+ uptake in permeabilized cardiac myocytes, but did not prevent the effect of protein kinase A. Anti-beta-receptor antibodies were present in 50% of the sera inhibiting Ca2+ uptake compared to 20% of those without inhibitory activity, (p less than 0.01). There was a strong correlation between the inhibition of sarcoplasmic reticulum Ca2+ transport and the HLA-DR4 phenotype (78% compared to 30% in patients with no inhibitory effect). These results suggest that immunological mechanisms play an important role in modifying sarcoplasmic reticulum function in about a third of the patients with detailed cardiomyopathy.
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PMID:Immune-mediated modulation of sarcoplasmic reticulum function in human dilated cardiomyopathy. 132 63

Antisera were produced in rabbits against synthetic peptides based on two regions of the cDNA sequence of the beta 1 subunit of bovine gamma-aminobutyric acidA (GABAA) receptors. The deduced amino acid sequences were similar in other beta subunits of bovine, rat, and chick receptors, predicting cross-reactability with all beta subunits. One antiserum (anti-beta e) was raised against an extracellular moiety near the invariant disulfide loop thought to be located near the neurotransmitter binding domain; the other (anti-beta c) was raised against an intracellular moiety containing a consensus sequence for cyclic AMP-dependent protein kinase phosphorylation of a serine residue. Predicted secondary structures suggested high potential immunogenicity for the chosen antigen peptides. Both antisera at high dilutions recognized the same polypeptide bands on western blots of GABAA receptors purified from three regions of bovine brain (four bands at 57, 54, 53, and 52 kDa in cerebral cortex) but fewer bands (57, 54, and 52 kDa) in hippocampus and cerebellum (one major band at 54 kDa, traces at 57 and 53 kDa). This is consistent with the presence of multiple beta subunits whose expression varies with brain region, as shown by molecular cloning. The anti-beta c antibody was able to immunoprecipitate purified GABAA receptor [3H]-muscimol binding, 87% in bovine cortex and 75% in total rat brain; the anti-beta e was unable to immunoprecipitate any antigen. These antibodies indicate a region-dependent heterogeneity of beta subunits and should be useful for analyzing structure, function, and localization of GABAA receptor subtypes in brain.
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PMID:Preparation of antibodies to beta subunits of gamma-aminobutyric acidA receptors. 132 21

A126-1B2 cells, a PKA (cAMP-dependent protein kinase)-deficient variant of PC12 cells, but not parental PC12 cells, form processes within 15-30 min of exposure to both nerve growth factor (NGF) and activators of protein kinase C when grown on tissue culture plastic (Glowacka and Wagner, J Neurosci Res 25: 453-462, 1990). Time-lapse microscopy has demonstrated that these processes are formed by a novel mechanism, i.e., rapid movement of the cell body away from a point of attachment, which morphologically resembles a growth cone. These "fast" neurites are attached to the substratum at a number of points, which display membrane activity in the form of active ruffling and the extension of filopodia and membrane pleats. Thus, these processes are formed by a mechanism distinct from that used by PC12 and other neuronal cells to form processes in culture. Wild-type PC12 cells also migrate and form fast neurites in response to a combination of NGF and phorbol 12-myristate 13-acetate (PMA), when they are grown in conditioned media or plates, suggesting that a secreted factor that can bind to the substratum is essential for the rapid formation of these neurites. Similarly, wild-type PC12 cells grown on a laminin-coated substratum also migrate and form "fast neurites" in response to a combination of NGF and PMA. This rapid migration is attenuated by an anti-alpha 1, beta 1-integrin antisera, implicating a laminin-integrin interaction; and it is inhibited by alpha-lactalbumin, suggesting an involvement of a beta 1,4 galactosyltransferase in the response. The formation of fast neurites is not dependent on concurrent protein synthesis, but it is inhibited by lithium, cytochalasin D, and methylthioadenosine or pretreatment of cells with NGF. Thus PC12 cells grown on the appropriate substrate have the ability to migrate rapidly and thereby form neuron-like processes within minutes of exposure to NGF and PMA. This morphological response to a combination of agents may provide an alternative means by which nerve cells form connections. Alternatively, it may reflect a mechanism that facilitates cellular migration during developmental processes.
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PMID:Synergistic effects of nerve growth factor and phorbol 12-myristate 13-acetate on rapid motility and process formation in PC12 cells: the role of laminin. 157 76

Heart rate and force can be increased by noradrenaline and adrenaline through an interaction with both beta 1-adrenoceptors (beta 1AR) and beta 2-adrenoceptors (beta 2 AR). Several ionic currents (I) can flow upon beta AR activation: ICa (through either beta 1AR or beta 2AR), INa, IK, and ICl. Calcium currents (ICa) can be increased directly by the alpha s unit of a GTP binding protein, Gs, or by coupling of Gs to adenylyl cyclase with subsequent formation of cyclic AMP, release of the catalytic unit of cyclic AMP-dependent protein kinase, and phosphorylation of calcium channels and other proteins. Chronic exposure (days or months), but not acute exposure (hours), to a catecholamine downregulates human heart beta 1AR. Acute desensitization partially uncouples human heart beta AR from the adenylyl cyclase. Both acute and chronic desensitization reduce positive inotropic responses to catecholamines. In human heart, catecholamine-induced activation of one beta 2AR causes the production of at least four times more cyclic AMP than activation of one beta 1AR. Chronic treatment of patients with beta 1AR-selective blockers paradoxically induces selective inotropic beta 2AR hyperresponsiveness, presumably by increasing coupling of beta 2AR to Gs. Several partial agonists with high affinity for heart beta 1AR and beta 2AR cause stimulant effects that are resistant to blockade of beta 1AR and beta 2AR. Such nonconventional partial agonists could perhaps interact with beta AR that resemble beta 3 adrenoceptors.
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PMID:Some aspects of heart beta adrenoceptor function. 165 75

Protein kinase c-beta-like immunoreactivity was studied in the adrenal gland of adult rats and at different pre- and postnatal stages of development (E17-P21) with an antibody specific to both the beta 1 and beta 2 subtypes of the kinase. In the adult rat adrenal gland, the immunoreactivity was seen in numerous nerve fibres in the adrenal medulla both in bundles and individually forming occasionally dense networks around chromaffin cell groups. Several protein kinase c-beta-immunoreactive fibres were also observed transversing the adrenal cortex towards the medulla. No remaining immunoreactive fibres two weeks after transection of the splanchnic nerve could be seen; nor was any immunoreactivity observed in the chromaffin cells of the adrenal medulla or in the cortical cells, but some faintly immunoreactive ganglion cells were detected in the adrenal medulla. The amount and distribution of protein kinase c-beta-like immunoreactivity in the fetal and developing adrenals was very similar to that seen in the adrenal glands of adult rats. On the basis of its localization, the beta-subtype of protein kinase c does not appear to be directly involved in the release of catecholamines from the adrenal medulla, but it might have a role in the regulation of neurotransmitter release from preganglionic cholinergic neurons.
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PMID:Protein kinase c-beta-like immunoreactivity in the developing and adult rat adrenal gland. 168 Aug 34


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