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)

The possible role of cyclic AMP-mediated phosphorylation events in the regulation of exocrine secretion after beta-adrenergic stimulation was examined in vitro in dispersed acinar cell aggregates from rat parotid gland. l-Isoproterenol, a beta-adrenergic agonist, stimulated endogenous activity of cyclic AMP-dependent protein kinase, alterations in the 32P content of 3 parotid phosphoproteins (increased 32P in 2, Mr = 27,000 and 14,000; decreased 32P in the remaining, Mr = 13,600), and amylase secretion in a dose-dependent manner. All responses were half-maximal within a range of l-isoproterenol concentrations of approximately 4 X 10(-8) to 5 X 10(-7) M. Examination of the time course of these 3 processes revealed that by 30 s after addition of l-isoproterenol, significant elevations in cyclic AMP-dependent protein kinase activity and alterations in the 32P content of the 3 parotid proteins had occurred, whereas secretion of amylase from cells was first detected 1-2 1/2 min after hormonal stimulation. Dibutyryl cyclic AMP (2 mM) elicited the same changes in parotid protein 32P content as l-isoproterenol. Our results support the concept of a role for cyclic AMP-regulated protein phosphorylation in the sequence of cellular events leading to exocrine protein secretion from the rat parotid gland following beta-adrenergic stimulation.
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PMID:beta-Adrenergic regulation of protein phosphorylation and its relationship to exocrine secretion in dispersed rat parotid gland acinar cells. 627 99

The effects of sodium nitroprusside, 8-bromo cyclic GMP, 8-bromoguanosine 5'-monophosphate, 8-bromo cyclic AMP, dibutyryl cyclic AMP, and isoproterenol on incorporation of (32)P into proteins in intact rat thoracic aorta were studied. Aortas were incubated in [(32)P]orthophosphate in order to label endogenous adenosine triphosphate. Agents were then added for various times and the tissues were homogenized and fractionated (100,000 x g for 60 min) into soluble and particulate fractions. Soluble and particulate fractions were subjected to isoelectric focusing followed by sodium dodecyl sulfate/polyacrylamide gel electrophoresis and autoradiographs were made. Nitroprusside induced a concentration-dependent increase in incorporation of (32)P into nine proteins and a decrease in (32)P incorporation into two proteins. Some of these proteins appeared in both the soluble and particulate fractions of homogenates; others appeared only in the soluble fraction. The pattern of (32)P incorporation was identical after 2- or 15-min exposure to nitroprusside and was mimicked by exposure to 50-500 muM 8-bromo cyclic GMP. 8-Bromoguanosine 5'-monophosphate did not alter (32)P incorporation. Dibutyryl cyclic AMP at 50 muM had no effect upon (32)P incorporation whereas a higher concentration (0.5 mM) caused increased or decreased (32)P incorporation into some, but not all, of the same proteins. 8-Bromo cyclic AMP (5 mM) produced only small changes in (32)P incorporation. The pattern of (32)P incorporation induced by a relatively high concentration of isoproterenol 0.1 mM was similar but not identical to that seen with 0.5 mM dibutyryl cyclic AMP. The present study indicates that the incorporation of (32)P into endogenous proteins of intact rat aorta can be regulated by nitroprusside. These effects can be mimicked by cyclic GMP analogues and only partially by cyclic AMP analogues or isoproterenol. Presumably, these effects of nitroprusside are mediated through a cyclic GMP-dependent process (protein kinase or phosphatase) which may play a role in the relaxant properties of nitroprusside and cyclic GMP.
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PMID:Sodium nitroprusside-induced protein phosphorylation in intact rat aorta is mimicked by 8-bromo cyclic GMP. 629 2

In order to determine if cyclic adenosine 3':5'-monophosphate- (cyclic AMP)-dependent protein kinase has a role in the expression of chemically induced differentiation of HL-60 cells, levels and subcellular distribution of this enzyme were studied during this process. Cyclic AMP binding protein and stimulated kinase activities increased moderately in cytosol and more markedly in nucleosol and nonhistone chromatin-associated protein fractions of cells induced to differentiate with dimethyl sulfoxide or 12-O-tetradecanoylphorbol-13-acetate. Retinoic acid induced similar cytosolic changes but less marked intranuclear increases. Nuclear increases did not occur in the differentiation-resistant subline, HL-60 Blast II, treated with dimethyl sulfoxide. DEAE-cellulose chromatography, as well as photoaffinity labeling and gel electrophoresis, disclosed higher ratios of type I to type II kinase in cytosol than in intranuclear fractions. Differences of the qualitative binding protein patterns between cytosol and nucleosol were enhanced following chemically induced differentiation. Dibutyryl cyclic AMP increased cytoplasmic and nuclear binding protein levels when given alone or in combination with retinoic acid or dimethyl sulfoxide, and it enhanced differentiation. These results suggest that intranuclear cyclic AMP-dependent protein kinase is associated with the expression of the differentiative program in HL-60 cells.
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PMID:Subcellular distribution of cyclic adenosine 3':5'-monophosphate-dependent protein kinase during the chemically induced differentiation of HL-60 cells. 632 33

We previously reported that the K562 cell line K562YO expressed a high level of the c-kit gene. In this study, we analyzed the mechanism of this expression and investigated the effects of the serine/threonine kinases such as protein kinase C (PKC) and cyclic adenosine 3',5'-monophosphate (cAMP)-dependent kinase (PKA) on it. The half-life of the c-kit mRNA in K562YO cells was greater than 10 hours, compared with 2 hours in the original K562 cells, which expressed a very low level of c-kit mRNA. This prolonged half-life can contribute to the high level of c-kit expression in K562YO cells. Cycloheximide (CHX), a protein synthesis inhibitor, caused increases in c-kit mRNA levels in K562YO cells. 12-O-tetradecanoylphorbol-13-acetate (TPA), by which PKC was activated at first and downregulated in a late phase, gradually decreased c-kit mRNA in K562YO cells until 9 hours and then returned to the control level 24 hours after treatment. TPA also rapidly decreased c-kit protein level on the membranes. In whole cells, c-kit protein was also decreased 6 hours after incubation with TPA. Calphostin C, a light-dependent PKC inhibitor, decreased c-kit mRNA levels within 30 minutes in a light-dependent manner. It also decreased c-kit protein in whole cells 2 hours after the addition. However, it increased the amount of c-kit protein on the cell surfaces. Dibutyryl cyclic AMP (dbc-AMP) increased c-kit mRNA as well as c-kit protein on membranes and in whole cells. Run-on transcriptional assay suggested that the agent (dbc-AMP) enhanced the transcription rate of the gene. These results suggest that c-kit protein on the membranes is downregulated by PKC activation and upregulated by PKC inhibition. In the whole cell lysate, c-kit proteins are decreased by PKC inhibition through downregulation of mRNA. On the other hand, the elevation of an intracellular cAMP level causes upregulation of both the mRNA and c-kit protein on membranes and in whole cells through enhanced transcription. Thus, c-kit gene expression is apparently modulated by PKC and PKA.
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PMID:High expression of c-kit in K562YO cells due to the prolonged half-life of its mRNA: the effects of modification with serine/threonine kinase signals. 753 32

To shed light on how axonal transport is regulated, we examined the possible roles of protein kinase A (PKA) in vivo suggested by our previous work (Sato-Yoshitake et al., 1992). Pharmacological probes or the purified catalytic subunit of PKA were applied to the permeabilized-reactivated model of crayfish walking leg giant axon, and the effect was monitored by the quantitative video-enhanced light microscopy and the quantitative electron microscopy. Dibutyryl cyclic AMP caused concentration-dependent transient reduction in the number of anterogradely transported small vesicles, while the retrogradely transported organelles and anterogradely transported mitochondria showed no decrease. This transient selective inhibition of anterograde vesicle transport was reversed by the application of a specific inhibitor of PKA (KT5720) in a concentration-dependent manner, and was reproduced by the application of the purified catalytic subunit of PKA and augmented by the application of adenosine 5'-O-(3-thiotriphosphate) (ATP gamma S). Corresponding biochemical study showed that several axoplasmic proteins including kinesin were specifically phosphorylated by the activation of the PKA pathway. These findings suggest the possible roles of PKA in the regulation of the direction of the axonal transport in vivo. The finding that only vesicle transport but not mitochondria transport was inhibited also suggests that the transport of vesicles and that of mitochondria are differently regulated and might be supported by different motors.
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PMID:The activation of protein kinase A pathway selectively inhibits anterograde axonal transport of vesicles but not mitochondria transport or retrograde transport in vivo. 753 26

Adrenergic regulation of RNA synthesis by in vivo stimulated parotid glands and dispersed parotid lobules was studied by a combination of in vivo and in vitro methods. Following a single intraperitoneal injection of isoproterenol, [3H]uridine incorporation into RNA was increased by 50% after the first hour. Amylase mRNA content was also elevated within 1 hr and was 2-3-fold higher than control values at 4 hr. An increase in the rate of total protein synthesis was detectable after 2 hr, and maximal rates were achieved 6 hr after isoproterenol administration. In dispersed parotid lobules, both isoproterenol and epinephrine stimulated [3H]uridine incorporation and at optimal concentrations increased incorporation by almost 200%. Phenylephrine (10 microM) caused a slight increase of about 20% whereas methoxamine (10 microM) had no effect. Stimulation by epinephrine was reversed by propranolol, but not by either phentolamine or prazosin. The increase in RNA synthesis induced by isoproterenol or epinephrine was dose dependent and half-maximal stimulation required 5.0 x 10(-8) M isoproterenol and 7.9 x 10(-7) M epinephrine. Dibutyryl cyclic AMP also stimulated [3H]uridine incorporation, whereas 8-bromo cyclic GMP, A23187 and phorbol myristate acetate had no effect. The importance of protein phosphorylation in mediating the observed stimulation was evaluated using protein kinase and phosphatase inhibitors. N-[2-(Methylamino)ethyl]-5-isoquinolinesulphonamide, an inhibitor of cyclic nucleotide-dependent protein kinases, substantially diminished the isoproterenol-induced stimulation. Okadaic acid treatment of lobules increased [3H]uridine incorporation. Furthermore, okadaic acid synergistically potentiated the stimulatory effect of a suboptimal concentration of isoproterenol. The results demonstrate that activation of the beta-adrenergic receptor induces the synthesis of certain RNA species in the parotid gland and that protein phosphorylation by a cyclic AMP-dependent protein kinase is a key event in the signal transduction pathway.
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PMID:Adrenergic regulation of RNA synthesis in the rat parotid gland. 768 14

AP-2 is a cell type-specific DNA-binding transcription factor that regulates selected target genes in vertebrate organisms. Here we investigated cell type-specific expression and regulation of AP-2 in neuroectodermal cell lineages. During retinoic acid (RA)-mediated differentiation of P19 embryonal carcinoma cells into neuroectodermal cell types that include immunohistochemically defined neurons and astrocytes, we observed a strong induction of AP-2 transcripts and protein. In contrast, AP-2 mRNA was not induced in P19 cells which undergo mesoendodermal differentiation in response to 1% dimethylsulfoxide or low concentrations of RA, respectively. The potential of both neurons and astrocytes to express AP-2 was ascertained by using cerebellar neurons and astrocytes derived from newborn mice. Unlike these types of cells, microglial cells do not express AP-2. Dibutyryl cyclic AMP further enhanced levels of AP-2 transcripts in both P19 astrocytes and primary astrocytes which also respond to agents elevating intracellular cAMP (noradrenaline, isoproterenol, forskolin). The cAMP-dependent induction of AP-2 could be blocked by inhibitors of protein kinase A. In contrast to its action in P19 cells, RA had no effect on AP-2 mRNA levels in primary astrocytes. Our results indicate that AP-2 may play a role as a retinoic acid-sensitive regulator during differentiation of neurons and glia from an embryonic neural precursor. Furthermore, AP-2 may be involved in gene transcription in both mature neurons and astrocytes.
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PMID:Cell type-specific regulation of expression of transcription factor AP-2 in neuroectodermal cells. 795 25

To assess the role of cAMP in the regulation of autophagy, we examined the effects of cAMP analogues and cAMP-elevating agents on freshly isolated rat hepatocytes, using electroinjected [3H]raffinose as an autophagy probe. Glucagon was found to stimulate, inhibit or have no effect on autophagy, depending on the inclusion of metabolites like pyruvate (which caused ATP depletion and autophagy suppression) and amino acids (a complete mixture that antagonized pyruvate) in the incubation medium. Inhibition was also observed with theophylline, a cAMP-elevating inhibitor of cyclic nucleotide phosphodiesterases, and with the adenylyl cyclase activator deacetylforskolin. At low concentrations of deacetylforskolin, the inhibition could be abolished by amino acids. N6,2'-O-Dibutyryladenosine 3',5'-monophosphate (Bt2-cAMP) strongly inhibited both autophagic sequestration of [3H]raffinose and overall autophagic protein degradation; again, amino acids abolished the autophagy-inhibitory effect of low Bt2-cAMP concentrations. Several other cAMP analogues (8-thiomethyl-cAMP, N6-benzoyl-cAMP, (S)-5,6-dichloro-1-D-ribofuranosylbenzimidazole 3',5'-[thio]monophosphate, (S)-8-bromoadenosine 3',5'-[thio]monophosphate) inhibited autophagy as well. The effect of Bt2-cAMP was rapid, dose-dependent, reversible and did not require concomitant protein synthesis. Neither Bt2-cAMP nor deacetylforskolin reduced intracellular ATP levels or cell viability, ruling out inhibition of autophagy by non-specific cytotoxicity. The autophagy-inhibitory effect of Bt2-cAMP could be substantially antagonized (40-50%) by KT-5720, a specific inhibitor of the cAMP-dependent protein kinase A, and by the nonspecific protein kinase inhibitor K-252a. Somewhat surprisingly, KN-62 and KT-5926, allegedly specific inhibitors of Ca2+/calmodulin-dependent protein kinase II and myosin light chain kinase, respectively, were also Bt2-cAMP-antagonistic. These results suggest that cAMP regulates the early, sequestrational step of hepatocytic autophagy by a highly conditional, dual mechanism, inhibition being predominant under most conditions in freshly isolated hepatocytes, whereas stimulation reportedly predominates in vivo. The effect of cAMP is probably mediated by protein kinase A, but other protein kinases would appear to participate in the regulation of autophagic sequestration as well.
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PMID:Role of cAMP in the regulation of hepatocytic autophagy. 861 61

1. The effects of the sequential stimulation of ciliary beat frequency (CBF) via two different phosphorylation cascades (dependent on protein kinase A (PKA) and calmodulin, respectively) were determined using video microscopy applied to a perfused preparation of human nasal respiratory epithelium in vitro. Dibutyryl cyclic AMP (db-cAMP) (10(-3) M) was used to stimulate PKA and the calcium ionophore 4-Br-A23187 (10(-5) M) was used to stimulate calmodulin-dependent phosphorylation. 2. Perfusion with db-cAMP (10(-3) M) alone showed an early rise in CBF (15.0 +/- 4%, mean +/- S.E.M., P < 0.05) by 10 min which remained elevated for 35 min; in contrast, the highest CBF response to 4-Br-A23187 (10(-5) M) alone was not achieved until 35 min (16.1 +/- 1.8%, P < 0.05). 3. When a db-cAMP stimulus was applied to cells which had been pre-incubated with 4-Br-A23187 for 30 min, a further rise in CBF (maximal at 20 min, 14.3 +/- 2%, P < 0.05) was observed. Reversing the sequence of perfusions, cells pre-incubated with db-cAMP showed no further rise in response to stimulation with 4-Br-A23187. 4. We hypothesized that PKA inhibited the response to the 4-Br-A23187. This notion was supported by the restoration of the CBF response (22.8 +/- 4%, P < 0.05) to 4-Br-A23187 when the cells were pre-incubated with the protein kinase inhibitor 1-(5-isoquinolinyl-sulphonyl)-2-methylpiperazine (10(-3) M), before the sequential perfusions with db-cAMP and 4-Br-A23187. We conclude that the A23187-dependent pathway, which regulates intrinsic CBF, is inhibited by db-cAMP but not vice versa.
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PMID:Asymmetric interactions between phosphorylation pathways regulating ciliary beat frequency in human nasal respiratory epithelium in vitro. 893 Aug 52

Human endothelial cells are injured by the action of leukocytes. We investigated the role of nitric oxide (NO) in the induction of injury to human pulmonary artery endothelial cells. NO has been a putative source of cytotoxic reactive oxygen species in some settings. Incubation of endothelial cells with neutrophils increased the release of lactate dehydrogenase activity and preloaded fura-2 from endothelial cells, indicating that neutrophils induce endothelial cell injury. This effect was augmented by treatment with carboxy-PTIO, which traps NO in the medium, or with L-NAME, an inhibitor of NO synthase. When endothelial cells were incubated with neutrophils stimulated by phorbol myristate acetate, an activator of protein kinase C, endothelial cell damage was further enhanced and the amount of NO in the medium was decreased. Dibutyryl cyclic AMP, a cell-permeable analogue of cyclic AMP, protected against neutrophil-induced endothelial cell injury and increased NO release into the medium. The effects of dibutyryl cyclic AMP were abrogated by treatment with H-89, a potent inhibitor of cyclic AMP-dependent protein kinase. The protective effect on neutrophil-induced endothelial cell injury by dibutyryl cyclic AMP was abolished by addition of carboxy-PTIO or L-NAME. Thus, our studies suggest that NO, presumably released from endothelial cells, protects against endothelial injury by activated neutrophils and the protective effect by cyclic AMP during coculture with activated neutrophils is mediated through the action of NO. However, when monocytes activated by lipopolysaccharide and IFN-gamma were used instead of neutrophils, endothelial cells were likewise injured, but a much higher level of NO was detected and injury was diminished by addition of carboxy-PTIO to the medium. These observations suggest that the high levels of NO released by activated monocytes contribute to endothelial injury, whereas low levels of NO protect endothelial cells against injury by neutrophils.
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PMID:The role of nitric oxide in human pulmonary artery endothelial cell injury mediated by neutrophils. 941 36


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