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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. Using internal perfusion and concentration-clamp procedures applied to Helix neurons, the effects of cAMP, Ca2+, and phorbol esters on ouabain-induced depression of acetylcholine Cl-dependent responses were determined. 2. Intracellular cAMP (10(-4) M) depressed those acetylcholine responses which were blocked by ouabain but had no effect on ouabain-insensitive acetylcholine responses. In the presence of elevated intracellular cAMP, ouabain had no further depressant effect on these acetylcholine responses. Both elevated cAMP and ouabain reduced the acetylcholine response without altering the current-voltage curves. 3. An increase in intracellular Ca2+ concentration depressed the amplitude of current induced by application of acetylcholine in neurons with ouabain-sensitive responses and shifted the dose-response relationship to the right. However, elevated Ca2+ did not reduce the maximal response induced by acetylcholine, nor did it prevent the reduction of that response by ouabain. 4. 12-O-Tetradecanoylphorbol-13-acetate (TPA), a potent stimulator of protein kinase C activity, caused depression of both the ouabain-sensitive and the ouabain-insensitive acetylcholine responses. The inhibitory effect of TPA was markedly enhanced after addition of ATP to the intracellular medium and was greatly reduced by cooling to 5 degrees C. The blocking effect of ouabain, however, reexamined in the presence of TPA. 5. These observations are consistent with the hypothesis that the depression of acetylcholine induced Cl--responses in Helix neurons is a result of an increase in intracellular cAMP concentration but is unrelated to activation of protein kinase C or increases in intracellular Ca2+.
Cell Mol Neurobiol 1992 Apr
PMID:The effects of cAMP, Ca2+, and phorbol esters on ouabain-induced depression of acetylcholine responses in Helix neurons. 131 66

HeLa cells attach to a variety of substrata but spread only on collagen or gelatin. Spreading is dependent on collagen-receptor upregulation, clustering, and binding to the cytoskeleton. This study examines whether second messengers are involved in initiating the spreading process on gelatin. The levels of cytosolic free calcium ([Ca++]i), cAMP, and cytoplasmic pH (pHi) do not change during cell attachment and spreading. However, a basal level of [Ca++]i and an alkaline pH(i) are required for spreading. There is an activation of protein kinase C (PKC) and a release of arachidonic acid (AA) on attachment and before cell spreading. Inhibition of PKC does not block cell spreading, indicating that PKC activation is not essential for spreading. Inhibition of phospholipase A2 blocks cell spreading, whereas addition of exogeneous AA overcomes this inhibitory effect. Among AA metabolic pathways, inhibitors of lipoxygenase (LOX) block cell spreading, suggesting that a LOX product(s) formed from AA initiates spreading. Clustering receptors for collagen with polyclonal antibodies, or with anti-collagen-receptor antigen-binding fragments (Fab) in combination with a secondary antibody, induce AA release. Also, AA is released when cells attach to either immobilized gelatin or immobilized Arg-Gly-Asp (RGD) peptide. Thus, AA is released whenever receptor clustering is observed. Receptor occupancy is not sufficient to release AA; when cells are treated with gelatin or RGD peptide in solution or anti-collagen-receptor Fab fragments without secondary antibody, conditions where receptor clustering is not observed, AA is not released. Thus, a LOX metabolite(s) of AA formed by collagen-receptor clustering is a second messenger(s) that initiates HeLa cell spreading. LOX inhibitors also block the spreading of bovine aortic endothelial cells, chicken embryo fibroblasts, and CV-1 fibroblasts on gelatin or fibronectin, indicating that other cells might use the same second messenger system in initiating cell-substratum adhesion.
Mol Biol Cell 1992 May
PMID:Spreading of HeLa cells on a collagen substratum requires a second messenger formed by the lipoxygenase metabolism of arachidonic acid released by collagen receptor clustering. 131 41

The effects of protein kinase C (PKC) activators on gamma-aminobutyric acidA (GABAA) receptor function were studied by two-electrode voltage-clamp in Xenopus oocytes expressing brain mRNA or subunit cDNAs and in isolated mouse brain cerebellar membrane vesicles (microsacs), using 36Cl- uptake. Both oocytes and microsacs showed transient (desensitizing) and sustained (nondesensitizing) GABAA receptor responses. In oocytes expressing brain mRNA, the PKC activator phorbol myristoyl acetate (PMA), but not the inactive analog phorbol 12-monomyristate, inhibited both transient and sustained GABA-gated chloride currents. The inhibition by PMA was concentration dependent, with an EC50 of approximately 5 nM, and resulted in a decrease in the efficacy, but not the potency, of GABA. Additionally, PMA inhibited GABA-gated chloride currents in oocytes expressing alpha 1 beta 1 gamma 2L subunit cDNAs. The effect of PMA on recombinant receptors was significantly antagonized by PKC inhibitory peptide (PKCI). In the microsac preparation, the PKC activators (-)-7-octylindolactam V and PMA inhibited the sustained phase of 36Cl- flux without altering the transient phase. The action of PMA was blocked by kinase inhibitors and by depletion of Mg-ATP and was mimicked by protein phosphatase inhibitors. These results demonstrate that activation of PKC inhibits GABAA receptor function, and the results from the microsac experiments suggest that PKC-dependent phosphorylation preferentially inactivates a nondesensitized form or state of the receptor.
Mol Pharmacol 1992 Jun
PMID:Activation of protein kinase C selectively inhibits the gamma-aminobutyric acidA receptor: role of desensitization. 131 47

The phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) is a potent activator of protein kinase C (PKC) and is known to affect a variety of biochemical processes in human breast cancer cells. In the present study we have employed MCF-7 cells to investigate the effects of TPA on inositol lipid signalling, the putative pathway leading to PKC activation and intracellular Ca2+ mobilization. Phosphoinositide hydrolysis in MCF-7 cells was stimulated by bombesin (BN) as evidenced by increases in both inositol phosphate production and cytidine diphosphate diacylglycerol (CDP-DG) accumulation. Pretreatment of MCF-7 cells with TPA caused attenuation of both these BN-induced responses. This inhibitory action of TPA on inositol phosphate production was mimicked by diacylglycerol analogues and was reversed by staurosporine, H-7 and tamoxifen, all known inhibitors of PKC. Furthermore, putative down-regulation of PKC by prolonged TPA pretreatment also reversed the inhibitory action of TPA and enhanced BN-induced phosphoinositide hydrolysis. TPA also inhibited BN-induced increases in cytosolic Ca2+ concentration ([Ca2+]i) and caused a dose-dependent inhibition of epidermal growth factor (EGF) binding in MCF-7 cells. However, EGF receptor occupancy was unaffected by BN. These data support an inhibitory role for PKC in the regulation of phosphoinositide hydrolysis and [Ca2+]i in breast cancer cells and provide a potential mechanism for feedback regulation of this signalling pathway in these cells.
Mol Cell Endocrinol 1992 Jun
PMID:Evidence for a role for protein kinase C in the modulation of bombesin-activated cellular signalling in human breast cancer cells. 132 70

The regulation of the guinea-pig pancreatic acinar plasma membrane Ca2+ pump by protein kinase A, protein kinase C and calmodulin was investigated. The results were compared with the effects of these regulators on the high affinity Ca(2+)-ATPase found in this membrane preparation. The catalytic subunit of cyclic AMP-dependent protein kinase stimulated Ca2+ transport 2-fold, but had no effect on Ca(2+)-dependent ATPase activity. Purified protein kinase C, the phorbol ester 12-O-tetradecanoyl phorbol-13-acetate and diacylglycerol derivative, 1-stearoyl-2-arachidonoyl-sn-glycerol, failed to stimulate the Ca(2+)-uptake but augmented the Ca(2+)-dependent ATPase activity. Exogenously added calmodulin failed to stimulate either activity. In addition, two antagonists of calmodulin activity, trifluoperazine and compound 48/80 produced a concentration-dependent inhibition of Ca(2+)-transport. These data suggest the presence of endogenous calmodulin within guinea-pig pancreatic acinar plasma membranes. Both calmodulin antagonists failed to influence the Ca(2+)-dependent ATPase activity. The ability of boiled extracts from guinea-pig pancreatic acinar plasma membranes to stimulate the Ca(2+)-ATPase activity in calmodulin-depleted erythrocyte plasma membranes confirmed the presence of endogenous calmodulin. Our results imply a role for calmodulin and cAMP-dependent protein kinase, but not protein kinase C, in the regulation of Ca2+ efflux from pancreatic acinar cells. These results also provide further evidence suggesting that the high affinity Ca(2+)-ATPase does not catalyze the plasma membrane Ca(2+)-transport activity observed in pancreatic acini.
Mol Cell Biochem 1992 Jun 26
PMID:Regulation of calcium transport in pancreatic acinar plasma membranes from guinea pig. 132 90

Microtubule-associated protein 2 (MAP2) kinase has been isolated and characterized from rat brain. The enzyme has an apparent M(r) of approximately 42,000 and its pI is 4.9. MAP2 was the preferred substrate, but it also phosphorylated myelin basic protein (MBP), histone V-S, tubulin and the PC12 protein substrate pp250. The enzyme is distinct from protein kinase C, cAMP-dependent kinase and the calcium/calmodulin-dependent kinases, as specific inhibitors of these kinases did not affect MAP2 phosphorylation. The addition of the relatively non-specific protein kinase inhibitor H7 (20 microM) had a modest inhibitory effect. The enzyme was active in both 5 mM Mn2+ and Mg2+, and displayed Kms for MAP2, MBP, and ATP of 56 nM, 254 nM, and 4 microM, respectively. This enzyme, which represents a low abundance protein in whole brain, is analogous to the MAP2 kinase observed in growth factor-stimulated cell lines.
Brain Res Mol Brain Res 1992 Jun
PMID:Isolation and characterization of microtubule-associated protein 2 (MAP2) kinase from rat brain. 132 16

Ovarian cytosol from pseudopregnant rats was heated to 80-90 degrees C for 2 min and precipitated proteins removed by centrifugation. The supernatant of the heated ovarian cytosol contained no protein kinase C activity but when added to a control preparation containing protein kinase C, enzyme activity was increased to 200% of control. The stimulatory activity was stable to heating for 10 min, was retained on a centrifugal filtration device with a 100,000 M(r) cut-off, did not affect cAMP-dependent protein kinase, was not extractable in petroleum ether or chloroform/methanol (2:1), and enhanced the phosphorylation of protein kinase C-specific peptide substrates. The stimulatory factor was calcium-dependent and could substitute for phosphatidylserine and diacylglycerol in the protein kinase C assay. This stimulatory factor may provide a mechanism whereby the response of protein kinase C to hormonal activation could be regulated by the cell.
Mol Cell Endocrinol 1992 Jul
PMID:Protein kinase C stimulatory activity in the pseudopregnant rat ovary. 132 55

The intracellular mechanisms of action of alpha-MSH in rat adrenocortical cells were examined. When rat adrenal capsule (largely glomerulosa) cells were stimulated with a range of concentrations of alpha-MSH there was significant stimulation of aldosterone secretion at 10(-10) mol/l, although cyclic AMP was not increased until high concentrations of alpha-MSH were used (10(-6) mol/l and above). However, cells incubated with ACTH showed an increase in aldosterone secretion at 10(-11) mol/l and levels of cyclic AMP were elevated at 10(-9) mol ACTH/l. When rat adrenal whole capsules were incubated with alpha-MSH, membrane-bound protein kinase C (PKC) activity was increased and cytosolic enzyme activity decreased, showing PKC activation. Stimulation with angiotensin II also induced translocation of PKC activity, but ACTH did not. When [3H]inositol-loaded glomerulosa cells were stimulated with alpha-MSH there was significant generation of [3H]inositol trisphosphate (IP3) at concentrations of alpha-MSH which stimulated secretion of aldosterone. Significantly increased levels of [3H]IP3 were also measured when loaded cells were exposed to angiotensin II. ACTH did not cause any significant stimulation of [3H]IP3 production at any concentration used. These results indicate that activation of PKC and phospholipase C is important in modulating the steroidogenic effect of alpha-MSH.
J Mol Endocrinol 1992 Aug
PMID:Studies on the intracellular mechanism of action of alpha-melanocyte-stimulating hormone on rat adrenal zona glomerulosa. 132 51

Primary fetal human adrenocortical cells of definitive zone origin were transfected by electroporation with pSV3neo, a plasmid coding for SV40 T antigen and neo, which confers resistance to the antibiotic G418. The clones obtained proliferated for 30 to 40 population doublings after isolation when grown under standard medium conditions, and then entered 'crisis'. When early-passage clones were incubated with cyclic AMP (1:1 N6-monobutyryl and 8-bromo analogues), cell rounding was observed, as in primary cultures of human adrenocortical cells. As previously shown in bovine adrenocortical cells, rounding was inhibited with a monoclonal antibody against urokinase plasminogen activator but not with a monoclonal antibody against tissue plasminogen activator. The regulation of the steroidogenic pathway in clones was investigated. The effects of cyclic AMP and activation of protein kinase C were examined in cells maintained in defined medium or in the presence of serum. 17 alpha-Hydroxylase was strongly induced by cyclic AMP, as evidenced by Northern blotting and by the conversion of progesterone or 25-hydroxy-[1,2-3H]cholesterol, this induction being blocked by low concentrations of 12-O-tetradecanoylphorbol-13-acetate (TPA). Cholesterol side-chain cleavage enzyme was strongly induced by cyclic AMP, and clones also showed low activities of 21-hydroxylase and 11 beta-hydroxylase. Under all circumstances levels of 3 beta-hydroxysteroid dehydrogenase (3 beta-HSD), as assessed by Northern blotting or by conversion of 25-hydroxycholesterol, were very low. 3 beta-HSD was not induced by cyclic AMP or TPA alone, but was induced by the combination of the two agents. The regulation of 17 alpha-hydroxylase and 3 beta-HSD resembles that previously described in primary cultures of human fetal adrenocortical cells. Thus, transfection with SV40 T antigen resulted in the production of clones which preserve the unique characteristics of the human adrenal cortex.
J Mol Endocrinol 1992 Aug
PMID:Expression of 17 alpha-hydroxylase and 3 beta-hydroxysteroid dehydrogenase in fetal human adrenocortical cells transfected with SV40 T antigen. 132 52

c-jun is a member of the family of immediate-early genes whose expression is induced by factors such as serum stimulation, phorbol ester, and differentiation signals. Here we show that increased Jun synthesis after serum stimulation is accompanied by a concomitant increase in phosphorylation. Several serine-threonine kinases were evaluated for their ability to phosphorylate Jun in vitro. p34cdc2, protein kinase C, casein kinase II, and pp44mapk phosphorylated Jun efficiently, whereas cyclic AMP-dependent protein kinase and glycogen synthase kinase III did not. The sites phosphorylated by p34cdc2 were similar to those phosphorylated in vivo after serum induction. The major sites of phosphorylation were mapped to serines 63, 73, and 246. Phosphorylation of full-length Jun with several kinases did not affect the DNA-binding activity of Jun homodimers or Fos-Jun heterodimers. Comparison of the DNA binding and in vitro transcription properties of wild-type and mutated proteins containing either alanine or aspartic acid residues in place of Ser-63, -73, and -246 revealed only minor differences among homodimeric complexes and no differences among Fos-Jun heterodimers. Thus, phosphorylation of Jun did not produce a significant change in dimerization, DNA-binding, or in vitro transcription activity. The regulatory role of phosphorylation in the modulation of Jun function is likely to be considerably more complex than previously suggested.
Mol Cell Biol 1992 Oct
PMID:Jun is phosphorylated by several protein kinases at the same sites that are modified in serum-stimulated fibroblasts. 132 60


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