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

In vitro, human B lymphocytes undergo long-term proliferation when activated through CD40, a protein expressed on their cell surface. The nature of CD40-dependent signals in proliferating fresh human Epstein-Barr virus-negative B lymphocytes is currently unknown. In this study, a CD40-dependent B cell culture system was used to examine the role of different signal transduction elements. Protein kinase C (PKC) depletion generated by a long-term phorbol 12 myristate 13-acetate treatment had weak effects on proliferation. Rather, tyrosine phosphorylation was shown to be directly involved in mediating CD40-dependent signals. The use of the protein tyrosine kinase (PTK)-specific inhibitor herbimycin A dramatically decreased cellular proliferation without altering the activity of the human immunodeficiency virus-1 long terminal repeat (HIV-1 LTR), a promoter largely dependent on the binding of nuclear factor kappa B (NF- kappa B). In contrast, the cAMP-dependent protein kinase specific inhibitor H-89 totally inhibited HIV-1 LTR activity at a concentration as low as 100 nM without affecting cellular proliferation. Electrophoretic mobility shift assay (EMSA) and supershift assay using an NF-kappa B binding sequence from the kappa light chain as a probe, revealed that both p65 (RelA) and c-Rel were present in CD40-stimulated B cells. While PKC depletion did not alter the NF-kappa B level, treatment of B lymphocytes with H-89 or herbimycin A provoked a decrease in the NF-kappa B level. These observations establish the importance of different signal transducing pathways leading to CD40 activation of B lymphocytes.
 ...
PMID:Tyrosine kinase and cAMP-dependent protein kinase activities in CD40-activated human B lymphocytes. 889 48

In the anterior pituitary, cGMP is produced in response to a number of stimuli, but intracellular events distal to cGMP production are obscure. Since cGMP-dependent protein kinase (PKG) is a major effector of cGMP actions in other tissues we have determined whether PKG and its specific substrates might be present and responsive to external signals in the ovine anterior pituitary. Photoaffinity labelling with [32P]cGMP revealed a specific 78 kDa protein in ovine anterior pituitary that comigrated with purified bovine lung PKG-I. PKG in protein extracts from anterior pituitary or cultured anterior pituitary cells was enriched by DEAE ion-exchange chromatography and assayed for activity. Both tissue and cultured cells had a relatively high PKG activity by comparison with aortic smooth muscle (known high activity) and brain (known low activity). Subcellular distribution studies showed that in anterior pituitary, aortic and brain, PKG activity was present in both cytosol and triton-extracted membrane fractions, while in platelets the activity was associated with only the membrane fraction. To determine if this PKG might be responsive to extracellular signals an activity ratio assay was used. Incubation of cultured cells with atrial natriuretic peptide (ANP) and sodium nitroprusside, activators of membrane and cytosolic guanylate cyclases respectively, increased the activity of PKG. To determine events distal to PKG activation, a search for potential substrates of PKG was performed. Few substrates were detectable upon addition of purified PKG to tissue lysates due to the high background activity of endogenous protein kinases in the anterior pituitary. However, 19 substrates of PKG were detected in heat-stable and 14 in acid-soluble protein extracts of the anterior pituitary, in which background phosphorylation was almost abolished. After partial purification through Q-Sepharose ion-exchange chromatography some of these proteins were preferentially phosphorylated by addition of PKG-I, while the others were additionally substrates of exogenous cAMP-dependent protein kinase (PKA) or Ca2+ and phospholipid-dependent protein kinase (PKC). A 132-kDa substrate showed an identical phosphopeptide map to a PKG substrate previously described in vascular smooth muscle and platelets. These data demonstrate for the first time the presence of functional PKG activity and multiple PKG substrates in the anterior pituitary where they may play a role in mediating the intracellular actions of cGMP.
 ...
PMID:Identification of cGMP-dependent protein kinase and its specific substrates in the anterior pituitary. 890 46

The phosphorylation and regulation of the URA7-encoded CTP synthetase (EC 6.3.4.2, UTP:ammonia ligase (ADP-forming)) from Saccharomyces cerevisiae by cAMP-dependent protein kinase (protein kinase A) were examined. Protein kinase A is the principal mediator of signals transmitted through the RAS/cAMP pathway in S. cerevisiae. The results of labeling experiments indicated that the phosphorylation of CTP synthetase was mediated by the RAS/cAMP pathway in vivo. In vitro, protein kinase A phosphorylated CTP synthetase at a serine residue with a stoichiometry consistent with one phosphorylation site per CTP synthetase subunit. Protein kinase A activity was dose- and time-dependent using CTP synthetase as a substrate. The dependence of protein kinase A activity on CTP synthetase was cooperative (n = 1.8) and the Km value for CTP synthetase was 73 nM. Phosphorylation of CTP synthetase with protein kinase A resulted in the stimulation (190%) of activity. The mechanism of this stimulation included an increase in the Vmax of the reaction with respect to UTP and ATP, a decrease in the Km for ATP, and a decrease in the cooperative kinetic behavior of the enzyme. Phosphorylated CTP synthetase was less sensitive to product inhibition by CTP. Protein kinase C also phosphorylates and activates CTP synthetase. Phosphorylation of CTP synthetase with protein kinases A and C together resulted in an increase in CTP synthetase activity that was slightly greater than that obtained when the enzyme was phosphorylated with either protein kinase alone.
 ...
PMID:Phosphorylation and regulation of CTP synthetase from Saccharomyces cerevisiae by protein kinase A. 891 May 20

Three-dimensional models of the five functional modules in human protein kinase C alpha (PKC alpha) have been generated on the basis of known related structures. The catalytic region at the C-terminus of the sequence and the N-terminal auto-inhibitory pseudo-substrate have been modeled using the crystal structure complex of cAMP-dependent protein kinase (cAPK) and PKI peptide. While the N-terminal helix of the catalytic region of PKC alpha is predicted to be in a different location compared with cAPK, the C-terminal extension is modeled like that in the cAPK. The predicted permissive phosphorylation site of PKC alpha, Thr 497, is found to be entirely consistent with the mutagenesis studies. Basic Lys and Arg residues in the pseudo-substrate make several specific interactions with acidic residues in the catalytic region and may interact with the permissive phosphorylation site. Models of the two zinc-binding modules of PKC alpha are based on nuclear magnetic resonance and crystal structures of such modules in other PKC isoforms while the calcium phospholipid binding module (C2) is based on the crystal structure of a repeating unit in synaptotagmin I. Phorbol ester binding regions in zinc-binding modules and the calcium binding region in the C2 domain are similar to those in the basis structures. A hypothetical model of the relative positions of all five modules has the putative lipid binding ends of the C2 and the two zinc-binding domains pointing in the same direction and may serve as a basis for further experiments.
 ...
PMID:Structural aspects of the functional modules in human protein kinase-C alpha deduced from comparative analyses. 891 29

Bone sialoprotein is a major noncollagenous protein of bone. Parathyroid hormone (PTH) was shown to cause a 2-4-fold increase in the steady-state levels of bone sialoprotein mRNAs within primary cultures of embryonic osteoblasts. The induction could be mimicked by both forskolin and phorbol 12-myristate 13-acetate and was not inhibited by cycloheximide. Transient expression of a approximately 1200-base pair avian 5' bsp promoter/reporter construct demonstrated similar inductions as mRNA levels. Co-transfection of an expression plasmid encoding heat-stable inhibitor of cAMP-dependent protein kinase, a peptide inhibitor of PKA, decreased both the basal and PTH-induced bsp transcription, while co-expression of the catalytic subunit of PKA-induced bsp expression 3-fold. Protein kinase C activation, on the other hand, did not appear to work through its activation of c-fos, since co-transfection of an expression clone for c-fos had no effect. Interestingly, heat-stable inhibitor of cAMP-dependent protein kinase also inhibited the phorbol 12-myristate 13-acetate induction, suggesting that the protein kinase C acts through some form of interaction with the cAMP/PKA pathway. A half-cAMP response element site in the bsp promoter was identified as the cis-acting element that mediated the PTH response by the transient transfections with reporter constructs containing nested deletions of the promoter or a heterologous promoter containing the cAMP response element. In conclusion, these data indicate that PTH stimulation of bsp gene expression is specific to osteoblasts and mediated by changing cellular cAMP/PKA levels. They further suggest that although protein kinase C is capable of stimulating the gene by itself, it plays a minimal role in mediating the PTH induction of bone sialoprotein.
 ...
PMID:Signal transduction pathways mediating parathyroid hormone stimulation of bone sialoprotein gene expression in osteoblasts. 893 23

The regulatory (R) domain of PKC alpha fused to glutathione-S-transferase (GST-R alpha) competitively inhibited PKC activity associated with extracts of Y1 mouse adrenocortical tumor cells and the activities of several specific PKC isozymes. GST-R alpha did not inhibit the activities of cAMP-dependent protein kinase, cGMP-dependent protein kinase or calmodulin-dependent myosin light chain kinase. GST-R alpha inhibited PKC activities 20 times more potently than did a synthetic peptide corresponding to the pseudosubstrate sequence of PKC alpha. In intact yeast cells, the R domain prevented PKC beta-1-induced inhibition of growth and cytokinesis. These results indicate that the R domain of PKC alpha acts as a specific, dominant inhibitor of PKC activity, and suggest that the PKC alpha R domain may provide a useful genetic tool to assess the roles of PKC in various signal transduction processes.
 ...
PMID:Molecular strategies for the dominant inhibition of protein kinase C. 896 21

A study is presented of the effect of the bile salt ursodeoxycholate (UDC) on protein phosphorylation by [gamma-32P]ATP in the cytosol from rat hepatocytes. Gel electrophoresis and corresponding autoradiograms of cytosolic proteins show that UDC promotes phosphorylation of at least eight different protein bands. Four of them (the 36, 60, 64 and 76 kDa) are phosphorylated by Ca2+ and phospholipid-dependent protein kinase (PKC); three (the 31, 51 and 71 kDa) are phosphorylated by cAMP-dependent protein kinase (PKA) and one protein band, with molecular weight of 34 kDa, apparently contains substrates of both PKC and PKA. Data are reported indicating that UDC can directly affect the intrinsic activity of protein kinases.
 ...
PMID:Ursodeoxycholate promotes protein phosphorylation in the cytosol of rat hepatocytes. 906 73

Protein phosphorylation is a primary means of mediating signal transduction events that control cellular processes. Accordingly, the activities of protein kinases and phosphatases are highly regulated. One level of regulation is that the subcellular distribution of several kinases and phosphatases is restricted by association with targeting proteins or subunits. This mechanism promotes rapid and preferential modulation of specific targets within a defined microenvironment in response to diffusible second messengers. The type II cAMP-dependent protein kinase (PKA) is targeted by association of its regulatory subunit (RII) with A-kinase anchoring proteins (AKAPs). To date, 36 unique AKAPs have been identified. Each of these proteins contains a conserved amphipathic helix responsible for AKAP association with cellular structures. Disruption of PKA/AKAP interaction with peptides patterned after the amphipathic helix region blocks certain cAMP responses, including the modulation of glutamate receptor ion-channel activity in neurons and transcription of cAMP-responsive genes. Yeast two-hybrid screening methods have identified neuronal specific AKAP79-binding proteins including the beta isoform of the phosphatase 2B, calcineurin. Biochemical and immunological studies have confirmed the two-hybrid results and identified additional members of this multienzyme signaling complex, including certain protein kinase C isoforms. These findings are consistent with colocalization of CaN, PKC, and type II PKA by AKAP79 and suggest a novel model for reversible phosphorylation in which the opposing kinase and phosphatase actions are colocalized in a signal transduction complex by association with a common anchor protein.
 ...
PMID:Dissection of protein kinase and phosphatase targeting interactions. 921 Feb 33

We investigated the modulation of the skeletal muscle L-type Ca2+ channel/dihydropyridine receptor in response to insulin-like growth factor-1 receptor (IGF-1R) activation in single extensor digitorum longus muscle fibers from adult C57BL/6 mice. The L-type Ca2+ channel activity in its dual role as a voltage sensor and a selective Ca2+-conducting pore was recorded in voltage-clamp conditions. Peak Ca2+ current amplitude consistently increased after exposure to 20 ng/ml IGF-1 (EC50 = 5.6 +/- 1.8 nM). Peak IGF-1 effect on current amplitude at -20 mV was 210 +/- 18% of the control. Ca2+ current potentiation resulted from a shift in 13 mV of the Ca2+ current-voltage relationship toward more negative potentials. The IGF-1-induced facilitation of the Ca2+ current was not associated with an effect on charge movement amplitude and/or voltage distribution. These phenomena suggest that the L-type Ca2+ channel structures involved in voltage sensing are not involved in the response to the growth factor. The modulatory effect of IGF-1 on L-type Ca2+ channel was blocked by tyrosine kinase and PKC inhibitors, but not by a cAMP-dependent protein kinase inhibitor. IGF-1-dependent phosphorylation of the L-type Ca2+ channel alpha1 subunit was demonstrated by incorporation of [gamma-32P]ATP to monolayers of adult fast-twitch skeletal muscles. IGF-1 induced phosphorylation of a protein at the 165 kDa band, corresponding to the L-type Ca2+ channel alpha1 subunit. These results show that the activation of the IGF-1R facilitates skeletal muscle L-type Ca2+ channel activity via a PKC-dependent phosphorylation mechanism.
 ...
PMID:Regulation of mouse skeletal muscle L-type Ca2+ channel by activation of the insulin-like growth factor-1 receptor. 927 27

The synaptic protein interaction (synprint) site on the N-type calcium channel alpha1B subunit binds to the soluble N-ethylmaleimide-sensitive attachment factor receptor (SNARE) proteins syntaxin and synaptosomal protein of 25 kDa (SNAP-25), and this association may be required for efficient fast synaptic transmission. Protein kinase C (PKC) and calcium and calmodulin-dependent protein kinase type II (CaM KII) phosphorylated a recombinant his-tagged synprint site polypeptide rapidly to a stoichiometry of 3-4 mol of phosphate/mol, whereas cAMP-dependent protein kinase (PKA) and cGMP-dependent protein kinase (PKG) phosphorylated the synprint peptide more slowly to a stoichiometry of <1 mol/mol. Two-dimensional phosphopeptide mapping revealed similar patterns of phosphorylation of synprint polypeptides and native rat brain N-type calcium channel alpha1B subunits by PKC and Cam KII. Phosphorylation of the synprint peptide with PKC or CaM KII, but not PKA or PKG, strongly inhibited binding of recombinant syntaxin or SNAP-25, even at a level of free calcium (15 microM) that stimulates maximal binding. In contrast, phosphorylation of syntaxin and SNAP-25 with PKC and CaM KII did not affect interactions with the synprint site. Binding assays with polypeptides representing the N- and C-terminal halves of the synprint site indicate that the PKC- and CaM KII-mediated inhibition of binding involves multiple, disperse phosphorylation sites. PKC or CaM KII phosphorylation of the synprint peptide also inhibited its interactions with native rat brain SNARE complexes containing syntaxin and SNAP-25. These results suggest that phosphorylation of the synprint site by PKC or CaM KII may serve as a biochemical switch for interactions between N-type calcium channels and SNARE protein complexes.
 ...
PMID:Phosphorylation of the synaptic protein interaction site on N-type calcium channels inhibits interactions with SNARE proteins. 927 28


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>