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 studied the in vitro effects of omega-3 fish oils and other fatty acids on the activity of crude protein kinase C from S49 lymphoma cells, on partially purified enzyme from rat cerebrum, on homogeneous protein kinase C from bovine brain, and, for comparison, on type I adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase. In the absence of exogenous phospholipid, the fish oils cis-5,8,11,14,17-eicosapentaenoic acid (EPA) and acid (DCHA) enhance the catalytic cis-4,7,10,13,16,19-docosahexaenoic activity of protein kinase C and support the binding of [3H]phorbol 12,13-dibutyrate, both to approximately 50% of the level supported by phosphatidylserine. In the presence of phosphatidylserine, the omega-3 fatty acids reduce catalytic activity and [3H]phorbol 12,13-dibutyrate binding by about one-half. The effects of the omega-3 fatty acids on enzyme activity suggest that fish oils act as partial agonists competitively with phosphatidylserine. EPA, DCHA, and arachidonate (but not a variety of saturated fatty acids) inhibit the cAMP-dependent protein kinase. Thus dietary fish oils and cellular fatty acids mobilized by the action of phospholipase A2 may differentially modulate the activities of protein kinase C and cAMP-dependent protein kinase. These data suggest means by which unsaturated fatty acids mobilized within cells may act as second messengers.
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PMID:Differential effects of omega-3 fish oils on protein kinase activities in vitro. 185 66

To determine the role of the stimulatory guanine nucleotide-binding protein, Gs, and adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase in the basal metabolism of beta-adrenergic receptors in S49 lymphoma cells, we measured the return of receptor number and function after irreversible blockade of receptors. After inactivation of receptors with the irreversible ligand N8-(bromoacetyl)-N'-[3-(4-indolyoxy)-2-hydroxypropyl]-(2)-1,8-diam ino-p- methane (BIM), beta-adrenergic receptors (defined as [125I]iodocyanopindolol binding sites) reappeared in a biphasic manner, the faster phase having a half-time (t 1/2) of 3-8 h (approximately 50% of the sites) and the slower phase greater than 40 h. Although the slow phase is not readily explained, recovery of binding sites during the first 10 h matched recovery of receptor function after BIM treatment (as measured by stimulation of cAMP accumulation) and recovery of receptor sites after downregulation induced by the agonist isoproterenol. Thus quantifying receptor recovery during the first 10 h after BIM treatment appears to be a reasonable method for examining basal receptor metabolism in S49 cells. Measured in this way, metabolism of beta-adrenergic receptors is very similar in wild-type S49 and the following variant clones: cyc- (absent Gs alpha), UNC and H21a (defective Gs alpha), and kin- (lacking cAMP-dependent protein kinase activity). Although previous data have demonstrated that agonist-promoted downregulation of beta-adrenergic receptors requires functional receptor-Gs coupling, the current data suggest that neither Gs nor cAMP-dependent protein kinase activity plays an important role in the regulation of basal metabolism of beta-adrenergic receptors.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Beta-adrenoceptor metabolism in wild-type, Gs, and protein kinase A-variant S49 cells. 197

The hormone-sensitive adenylyl cyclase system is under dual control, receiving both stimulatory and inhibitory inputs. Guanine nucleotide-binding regulatory proteins (G-proteins) transduce signals from cell surface receptors to effectors such as adenylyl cyclase. Hormonal stimulation is propagated via Gs, inhibition by Gi. Persistent (24-h) activation of the stimulatory pathway of adenylyl cyclase by the diterpene forskolin or the beta-adrenergic agonist isoproterenol in S49 mouse lymphoma cells enhanced the effects of somatostatin mediated via the inhibitory pathway of adenylyl cyclase. Stimulating cells with forskolin or isoproterenol for 24 h resulted in a 3-fold increase in the steady-state levels of Gi alpha 2 and a 25% decline in Gs alpha, as quantified by immunoblotting. Within 12 h of stimulation of adenylyl cyclase, Gi alpha 2 mRNA levels increased 4-fold, measured by DNA-excess solution hybridization. Gs alpha mRNA levels, in contrast, increased initially (25%), but then declined to 75% of control. In S49 variants that lack functional protein kinase A (kin-), stimulation by isoproterenol failed to alter Gi alpha 2 expression at either the protein or the mRNA levels. A 3-fold increase in relative synthesis rate and no change in the half-life (approximately 80 h) of Gi alpha 2 was observed in response to forskolin stimulation. Although Gs alpha synthesis increased (70%) modestly in response to forskolin stimulation, the half-life of Gs alpha actually decreased from 55 h in naive cells to 34 h in treated cells. Thus, the two G-protein-mediated pathways controlling adenylyl cyclase display "cross-regulation." Persistent activation of the stimulatory pathway increases Gi alpha 2 mRNA and expression. Transiently elevated Gs alpha mRNA levels are counterbalanced by a reduction in the half-life of the protein.
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PMID:Cross-regulation between G-protein-mediated pathways. Stimulation of adenylyl cyclase increases expression of the inhibitory G-protein, Gi alpha 2. 211 18

Ribonucleotide reductase activity in S49 T lymphoma cells is cell cycle regulated by de novo protein synthesis of the M2 subunit. There is maximal enzyme activity in S and G2/M phase with low activity and low concentrations of the M2 subunit in G1 phase. Pharmacologic concentrations of cyclic AMP arrest S49 cells in the G1 phase of the cell cycle. We investigated the effect of cyclic AMP on M2 messenger RNA concentrations using RNA from exponentially growing and elutriated, cell cycle-enriched populations. To discern whether cyclic AMP-induced G1 arrest was associated with low concentrations of M2-specific messenger RNA, we probed blots with a full-length cDNA for M2. Cell cycle variation in M2 messenger RNA concentrations was similar in wild-type, hydroxyurea-resistant cells with amplified M2 activity, and cyclic AMP-dependent protein kinase-deficient cell lines. All lines had low amounts of M2-specific mRNA in early G1, an increase at the late G1/early S phase interface, a decrease in mid S phase, and another increase in late S phase that continued through G2/M. These concentrations did not directly correlate with enzyme activity, suggesting other regulatory effects might participate in determining ribonucleotide reductase activity. Cyclic AMP exposure appeared to induce cell cycle arrest in early G1 with low M2-specific messenger RNA concentration. This effect reversed upon washout of the cyclic AMP and was dependent on functional cyclic AMP-dependent protein kinase (PKA). These results suggest that cyclic AMP arrests S49 mouse T lymphoma cells in early G1 prior to transcriptional activation of the M2 gene.
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PMID:Effect of cyclic AMP on the cell cycle regulation of ribonucleotide reductase M2 subunit messenger RNA concentrations in wild-type and mutant S49 T lymphoma cells. 215 14

S49 wild-type mouse lymphoma cells grown in 3 nM epinephrine are extensively desensitized. Cellular cAMP responses to subsequent challenges with 100 nM epinephrine are reduced by as much as 80-90%. In this report, we document that protein kinase activity ratios were also attenuated. For example, the activity ratios in naive cells were increased from 0.26 +/- 0.02 to 0.72 +/- 0.04 by incubation with 100 nM epinephrine for 2 min, whereas in cells grown in 3 nM epinephrine for 24 hr before the experiment they were 0.19 +/- 0.02 and 0.29 +/- 0.03. Attenuated protein kinase activity ratios were obvious at epinephrine challenge concentrations ranging from 10 to 1000 nM. Three kinds of experiments provided evidence that the reduced ratios in desensitized cells were secondary to diminished cAMP responses rather than to changes in the cAMP-dependent protein kinase itself. Firstly, when protein kinase activity ratios were plotted against cAMP levels in naive and desensitized cells, the points fell along a common line. Secondly, cell-free cAMP-dependent protein kinase preparations from naive or epinephrine-treated cells had similar activities in the presence of maximal exogenous cAMP and similar half-maximal cAMP concentrations. Finally, the levels of cAMP-binding proteins in extracts from naive and desensitized cells were essentially identical. We conclude that desensitization of S49 cells by very low levels of epinephrine significantly reduced cAMP-dependent protein kinase responses to much higher concentrations of the catecholamine.
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PMID:cAMP-dependent protein kinase responses of S49 cells are reduced by growth in low epinephrine concentrations. 216 19

5'-p-Fluorosulfonylbenzoyladenosine (FSBA) is a useful reagent for the affinity labeling of adenine nucleotide binding proteins. We have developed an immunochemical approach to the detection of proteins that have been covalently modified with FSBA, which provides an alternative to the use of a radiolabeled ligand. Antibodies have been prepared against FSBA-modified glutamate dehydrogenase and purified by chromatography on ATP-agarose. The resulting affinity-purified antibodies react on Western blots only with proteins that have been labeled previously with the affinity reagent. The degree of immunoreactivity on Western blots correlates well with the extent of covalent modification as shown by studies on the modification and inhibition of the catalytic subunit of cAMP-dependent protein kinase. In crude cellular extracts, numerous proteins can be labeled with FSBA and then detected by using this approach. The labeling and subsequent detection of these proteins can be blocked by including an excess of MgATP, which competes with FSBA for nucleotide-binding sites. The labeling of specific proteins in crude mixtures is saturable, as shown by labeling studies of p56lck, a protein-tyrosine kinase that is abundantly expressed in membranes from the T lymphoma cell line LSTRA.
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PMID:Immunochemical detection of adenine nucleotide-binding proteins with antibodies to 5'-p-fluorosulfonylbenzoyladenosine. 228 46

The S49 mouse lymphoma mutant cell line Kin- is resistant to the cytotoxic effects of elevated cAMP levels, has no detectable cAMP-dependent protein kinase activity, and has depressed levels of cAMP-binding regulatory subunits. We demonstrate that although the Kin- cell line lacks detectable catalytic subunit protein, these cells express wild-type levels of mRNA for both C alpha and C beta catalytic subunit isoforms. Translation of C alpha mRNA appears to be normal in the Kin- cell, based on the observation that C alpha mRNA associates with large polyribosomes in both wild-type and Kin- cells. We cloned the C alpha cDNA from Kin- cells and show that its transient expression in another cell type leads to activation of a cAMP-sensitive luciferase reporter gene, suggesting that functional C alpha protein is made. In addition to having catalytic activity, the C alpha subunit from Kin- cells is inhibited in the presence of mouse RI alpha regulatory subunit, indicating that formation of the holoenzyme complex is normal. We suggest that the mutation responsible for the Kin- phenotype is in a cellular component that directly or indirectly causes Kin- catalytic subunit protein to be degraded rapidly.
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PMID:The S49 Kin- cell line transcribes and translates a functional mRNA coding for the catalytic subunit of cAMP-dependent protein kinase. 230 38

ECH408-1 is a murine B cell lymphoma expressing idiotypically and allotypically distinguishable transfected and endogenous IgD. Previously, we demonstrated that this cell line was not growth inhibited by antibodies directed at membrane IgD, but could be inhibited by antibodies which crosslink membrane IgM. Herein, we demonstrate that both anti-mu and anti-delta will cause calcium mobilization in this transfected cell line; this is followed by a period during which antibodies against the alternative isotype are unable to induce significant increases in intracellular calcium concentrations. This phenomenon, called "desensitization," is short-lived, lasting 20 min. We further demonstrate that acute desensitization of these cells by anti-delta has no effect on immediate growth inhibition which is elicited by anti-mu. These data confirm our earlier proposal that the rapid, initial calcium response seen in these lymphomas is not required for the negative signal for growth. Moreover, we also demonstrate that pretreatment of these lymphoma cells with phorbol myristate acetate (PMA) also renders these lymphoma cells temporarily incapable of manifesting a significant calcium signal. Nonetheless, PMA-pretreated B lymphoma cells are not altered in their subsequent sensitivity to anti-mu growth inhibition, nor are they affected in their resistance to inhibition by anti-delta. Our data confirm the proposal that neither the calcium signal nor protein kinase-C activation is involved in the modulation of B lymphoma growth.
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PMID:Lymphoma models for B cell activation and tolerance. VIII. Cross-desensitization by sIgM and sIgD and its effects on growth regulation by anti-isotype antibodies. 232 38

Expression of cellular src-gene product (p60c-src) in human leukemia-lymphoma cell lines was analysed by flow cytometry using a monoclonal antibody (McAb), H2B4 which recognizes p60c-src protein in human cells. In several human leukemia-lymphoma cell lines (K562, Namalva, HL60, U937), p60c-src expression was higher than in peripheral mononuclear cells from healthy volunteers. Some non-lymphoid leukemia cells can be induced to differentiate into monocyte-macrophages by 12-O-tetradecanoyl phorbol-13-acetate (TPA). K562 cells were also induced to differentiate not only morphologically but also functionally into monocyte-macrophages by TPA. Flow cytometric analyses using the McAb H2B4 revealed that the amount of p60c-src expression in K562 cells markedly decreased during TPA induced differentiation. The activity of protein kinase associated with p60c-src in K562 cells was determined employing IgG of immunized rabbit serum specific for p60c-src. The immunized rabbit IgG heavy chain phosphorylation by protein kinase also decreased after the induced differentiation. We detected p60c-src protein in acute lymphoid leukemia cells as well as acute non-lymphoid leukemia cells freshly isolated from patients. The amount of p60c-src protein decreased in some acute non-lymphoid leukemia cases, but it increased in others after TPA induced differentiation. No correlation was observed between FAB classification of acute leukemias and the amount of endogenous p60c-src expression.
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PMID:Alteration of p60c-src expression in human leukemia-lymphoma cells correlated with induced differentiation. 245 97

Cyclic AMP arrests T lymphocytes in the G1 phase of the cell cycle, and prolonged exposure results in cytolysis. Both of these effects require cyclic AMP-dependent protein kinase. We recently observed that some S49 mouse T lymphoma cell lines selected for hydroxyurea resistance were not arrested in G1 by cyclic AMP. Further analysis revealed that these cell lines were cyclic AMP-dependent protein kinase deficient, and conversely, other cyclic AMP-dependent protein kinase deficient cell lines not selected for hydroxyurea resistance were two- to threefold more hydroxyurea resistant. However, hydroxyurea is a specific inhibitor of ribonucleotide reductase and does not inhibit this kinase. We subsequently showed that cyclic AMP-dependent protein kinase will phosphorylate the M2 but not the M1 subunit of ribonucleotide reductase in vitro, and this phosphorylation will diminish CDP reductase activity. In vivo phosphorylation of M2 occurred under conditions similar to those that generate cell cycle arrest. We conclude that the M2 subunit of ribonucleotide reductase can be a target of cyclic AMP-dependent protein kinase. The phosphorylated enzyme has diminished activity, and this may play a role in cyclic AMP-induced lymphocyte cell cycle arrest.
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PMID:M2 subunit of ribonucleotide reductase is a target of cyclic AMP-dependent protein kinase. 253 34


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