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)

Phospholamban is a negative regulator of the sarcoplasmic reticulum Ca(2+)-pumping ATPase. Phosphorylation of phospholamban activates the ATPase and decreases the level of cytosolic calcium. Phospholamban is phosphorylated in heart by cAMP-dependent protein kinase, cGMP-dependent protein kinase and calcium/calmodulin-dependent protein kinase II (CM-kinase-II) and in smooth muscle cells by cGMP-dependent protein kinase. In contrast to heart muscle, phospholamban is poorly phosphorylated by CM-kinase-II in extracts of rat aortic smooth muscle cells. Rat aorta phospholamban amino acid sequence was identical to dog heart. The peptide substrate specificity of CM-kinase-II from rat aorta was the same as that from rat heart. The lack of phosphorylation of rat aorta phospholamban by the CM-kinase-II appears to result from the relatively low abundance of phospholamban in smooth muscle.
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PMID:Phosphorylation of phospholamban in aortic smooth muscle cells and heart by calcium/calmodulin-dependent protein kinase II. 785 66

Multiple processes lead to neuronal death after ischemia, but the generation of nitric oxide (NO) is a key component in this cascade of events. The mechanisms that regulate the extent of neuronal degeneration during anoxia and NO toxicity are multifactorial. Neuronal death may be modulated by the activity of signal transduction systems that influence the toxicity of NO or its metabolic products such as cGMP. The enzyme responsible for the production of NO, nitric oxide synthase (NOS), is phosphorylated by protein kinase C (PKC), the cAMP-dependent protein kinase (PKA), and the calcium/calmodulin-dependent protein kinase II (CaM-II). We examined in primary cultured hippocampal neurons whether the protein kinases PKC, PKA, CaM-II, and cGMP-dependent protein kinase modified the toxic effects of anoxia and NO. Down-regulation of PKC activity with PMA (1 microM) increased hippocampal neuronal survival during anoxia and NO exposure from approximately 22% to 88%. Inhibitors of PKC activity (H-7, H-8, sphingosine, and staurosporine) also were neuroprotective. Down-regulation of PKC activity increased survival during anoxia even in the presence of the NOS inhibitor, N omega-methyl-L-arginine. Thus, although down-regulation of PKC activity may increase neuronal survival by decreasing NOS activity, it also is likely that PKC contributes to ischemic neuronal death by mechanisms that are independent of NOS. Inhibition of the cGMP-dependent protein kinase activity, but not the activity of the CaM-II also was neuroprotective during NO administration. In contrast to the protective effects of inhibition of PKC and the cGMP-dependent protein kinase, activation rather than inhibition of PKA increased hippocampal neuronal survival during NO exposure. These results indicate that neuronal survival during anoxia and NO exposure is linked to the modulation of PKC, PKA, and cGMP-dependent protein kinase activity but is not dependent on the CaM-II pathway. Understanding the involvement of PKC, PKA, and the cGMP-dependent protein kinase in modulating the effect of neuronal death during ischemia and NO toxicity may help in directing future therapeutic modalities for cerebrovascular disease.
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PMID:Protein kinases modulate the sensitivity of hippocampal neurons to nitric oxide toxicity and anoxia. 823 Mar 23

The neuronal protein neurogranin, also known as RC3, is a selective substrate for protein kinase C (PKC). We synthesized a peptide corresponding to the phosphorylation domain of neurogranin (amino acids 28-43) and characterized its properties as a PKC substrate. Neurogranin(28-43) was phosphorylated by purified PKC with a Km of 150 nM. No significant phosphorylation of the peptide by either cAMP-dependent protein kinase or by calcium/calmodulin-dependent protein kinase II could be detected. Thus, neurogranin(28-43) is a potent and selective substrate for PKC. We tested several peptide analogues of neurogranin(28-43) for their substrate potency and specificity as kinase substrates, in order to help elucidate the structural determinants involved in the phosphorylation of substrates by PKC. Substituting Arg36 with Ile caused a significant reduction in the affinity for PKC. Replacing Lys30 with Arg enhanced the catalytic efficiency (Vmax/Km) for PKC but diminished the selectivity of the substrate for PKC. These results support the generally held model that basic amino acids on both sides of the phosphorylated Ser are important structural determinants in PKC substrates. However, the data also suggest that the presence of particular basic amino acids (Arg vs Lys) can contribute to the degree of selectivity of a substrate for PKC. Replacement with Ala of Phe35, the amino acid adjacent to the Ser34 phosphorylation site, resulted in a peptide with greatly diminished potency as a PKC substrate. This finding indicates a critical role of Phe35 in modulating binding and phosphorylation of neurogranin-derived peptides by PKC.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Studies with synthetic peptide substrates derived from the neuronal protein neurogranin reveal structural determinants of potency and selectivity for protein kinase C. 842 32

We analyzed whether synaptic membrane trafficking proteins are substrates for casein kinase II, calcium/calmodulin-dependent protein kinase II, and cAMP-dependent protein kinase (PKA), three kinases implicated in the modulation of synaptic transmission. Each kinase phosphorylates a specific set of the vesicle proteins syntaxin 1A, N-ethylmaleimide-sensitive factor (NSF), vesicle-associated membrane protein (VAMP), synaptosome-associated 25-kDa protein (SNAP-25), n-sec1, alpha soluble NSF attachment protein (alpha SNAP), and synaptotagmin. VAMP is phosphorylated by calcium/calmodulin-dependent protein kinase II on serine 61. alpha SNAP is phosphorylated by PKA; however, the beta SNAP isoform is phosphorylated only 20% as efficiently. alpha SNAP phosphorylated by PKA binds to the core docking and fusion complex 10 times weaker than the dephosphorylated form. These studies provide a first glimpse at regulatory events that may be important in modulating neurotransmitter release during learning and memory.
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PMID:Phosphorylation of synaptic vesicle proteins: modulation of the alpha SNAP interaction with the core complex. 887 42

Protein kinases play an important role in controlling synaptic strength at excitatory synapses on CA1 pyramidal cells. We examined the effects of activating cAMP-dependent protein kinase or protein kinase C (PKC) on the frequency and amplitude of miniature excitatory postsynaptic currents (mEPSCs) with perforated patch recording techniques. Both forskolin and phorbol-12,13-dibutryate (PDBu) caused a large increase in mEPSC frequency, but only PDBu increased mEPSC amplitude, an effect that was not observed when standard whole cell recording was performed. These results support biochemical observations indicating that PKC, similar to calcium/calmodulin-dependent protein kinase II, has an important role in controlling synaptic strength via modulation of AMPA receptor function, potentially through the direct phosphorylation of the GluR1 subunit.
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PMID:Effects of PKA and PKC on miniature excitatory postsynaptic currents in CA1 pyramidal cells. 981 84

Brief bath application of N-methyl-D-aspartate (NMDA) to hippocampal slices produces long-term synaptic depression (LTD) in CA1 that is (1) sensitive to postnatal age, (2) saturable, (3) induced postsynaptically, (4) reversible, and (5) not associated with a change in paired pulse facilitation. Chemically induced LTD (Chem-LTD) and homosynaptic LTD are mutually occluding, suggesting a common expression mechanism. Using phosphorylation site-specific antibodies, we found that induction of chem-LTD produces a persistent dephosphorylation of the GluR1 subunit of AMPA receptors at serine 845, a cAMP-dependent protein kinase (PKA) substrate, but not at serine 831, a substrate of protein kinase C (PKC) and calcium/calmodulin-dependent protein kinase II (CaMKII). These results suggest that dephosphorylation of AMPA receptors is an expression mechanism for LTD and indicate an unexpected role of PKA in the postsynaptic modulation of excitatory synaptic transmission.
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PMID:NMDA induces long-term synaptic depression and dephosphorylation of the GluR1 subunit of AMPA receptors in hippocampus. 985 70

Rats were trained in one-trial step-down inhibitory avoidance and tested either 3 h or 31 days later. Ten minutes prior to the retention test, through indwelling cannulae placed in the CA1 region of the dorsal hippocampus, they received 0.5 microl infusions of: saline, a vehicle (2% dimethylsulfoxide in saline), the glutamate NMDA receptor blocker, aminophosphonopentanoic acid (AP5) (5.0 microg), the AMPA/kainate receptor blocker, cyanonitroquinoxaline dione (CNQX) (0.25 or 1.25 microg), the metabotropic receptor antagonist, methylcarboxyphenylglycine (MCPG) (0.5 or 2.5 microg), the inhibitor of calcium/calmodulin-dependent protein kinase II (KN62) (3.5 microg), the inhibitor of cAMP-dependent protein kinase (PKA), Rp-cAMPs (0.1 or 0.5 microg), the stimulant of the same enzyme, Sp-cAMPs (0.1 or 0.5 microg), or the inhibitor of the mitogen-activated protein kinase (MAPK) kinase, PD098059 (10 or 50 microM). CNQX, KN62 and PD098059 were dissolved in the vehicle; the other drugs were dissolved in saline. All these drugs, at the same doses, had been previously found to affect short- and long-term memory formation of this task. Retrieval measured 3 h after training (short-term memory) was blocked by CNQX and MCPG, and was unaffected by all the other drugs. In contrast, retrieval measured at 31 days was blocked by MCPG, Rp-cAMPs and PD098059, enhanced by Sp-cAMPs, and unaffected by CNQX, AP5 or KN62. The results indicate that, in CA1, glutamate metabotropic receptors are necessary for the retrieval of both short- and long-term memory; AMPA/kainate receptors are necessary for short-term but not long-term memory retrieval, and NMDA receptors are uninvolved in retrieval. Both the PKA and MAPK signalling pathways are required for the retrieval of long-term but not short-term memory.
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PMID:Different hippocampal molecular requirements for short- and long-term retrieval of one-trial avoidance learning. 1084 Jan 35

Choline acetyltransferase synthesizes acetylcholine in cholinergic neurons and, in humans, may be produced in 82- and 69-kDa forms. In this study, recombinant choline acetyltransferase from baculovirus and bacterial expression systems was used to identify protein isoforms by two-dimensional SDS/PAGE and as substrate for protein kinases. Whereas hexa-histidine-tagged 82- and 69-kDa enzymes did not resolve as individual isoforms on two-dimensional gels, separation of wild-type choline acetyltransferase expressed in insect cells revealed at least nine isoforms for the 69-kDa enzyme and at least six isoforms for the 82-kDa enzyme. Non-phosphorylated wild-type choline acetyltransferase expressed in Escherichia coli yielded six (69 kDa) and four isoforms (82 kDa) respectively. Immunofluorescent labelling of insect cells expressing enzyme showed differential subcellular localization with the 69-kDa enzyme localized adjacent to plasma membrane and the 82-kDa enzyme being cytoplasmic at 24 h. By 64 h, the 69-kDa form was in cytoplasm and the 82-kDa form was only present in nucleus. Studies in vitro showed that recombinant 69-kDa enzyme was a substrate for protein kinase C (PKC), casein kinase II (CK2) and alpha-calcium/calmodulin-dependent protein kinase II (alpha-CaM kinase), but not for cAMP-dependent protein kinase (PKA); phosphorylation by PKC and CK2 enhanced enzyme activity. The 82-kDa enzyme was a substrate for PKC and CK2 but not for PKA or alpha-CaM kinase, with only PKC yielding increased enzyme activity. Dephosphorylation of both forms of enzyme by alkaline phosphatase decreased enzymic activity. These studies are of functional significance as they report for the first time that phosphorylation enhances choline acetyltransferase catalytic activity.
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PMID:Expression, purification and characterization of recombinant human choline acetyltransferase: phosphorylation of the enzyme regulates catalytic activity. 1086 Dec 22

Reversal of long term potentiation (LTP) may function to increase the flexibility and storage capacity of neuronal circuits; however, the underlying mechanisms remain incompletely understood. We show that depotentiation induced by low frequency stimulation (LFS) (2 Hz, 10 min, 1200 pulses) was input-specific and dependent on N-methyl-d-aspartate (NMDA) receptor activation. The ability of LFS to reverse LTP was mimicked by a brief application of NMDA. This NMDA-induced depotentiation was blocked by adenosine A(1) receptor antagonist. However, the reversal of LTP by LFS was unaffected by metabotropic glutamate receptor antagonism. This LFS-induced depotentiation was specifically prevented by protein phosphatase (PP)1 inhibitors, okadaic acid, and calyculin A but not by the PP2A or PP2B inhibitors. Furthermore, by using phosphorylation site-specific antibodies, we found that LFS-induced depotentiation is associated with a persistent dephosphorylation of the GluR1 subunit of amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor at serine 831, a protein kinase C and calcium/calmodulin-dependent protein kinase II (CaMKII) substrate, but not at serine 845, a substrate of cAMP-dependent protein kinase. This effect was mimicked by bath-applied adenosine or NMDA and was specifically prevented by okadaic acid. Also, the increased phosphorylation of CaMKII at threonine 286 and the decreased PP activity seen with LTP were overcome by LFS, adenosine, or NMDA application. These results suggest that LFS erases LTP through an NMDA receptor-mediated activation of PP1 to dephosphorylate amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors and CaMKII in the CA1 region of the hippocampus.
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PMID:Characterization of the mechanism underlying the reversal of long term potentiation by low frequency stimulation at hippocampal CA1 synapses. 1167 81

Glucagon like peptide-1 (GLP1) is a G(s)-coupled receptor agonist that exerts multiple effects on pancreatic beta-cells, including the stimulation of insulin gene expression and secretion. In this report, we show that treatment of the mouse pancreatic beta-cell line MIN6 with GLP1 leads to the glucose-dependent activation of Erk. These effects are mimicked by forskolin, a direct activator of adenylate cyclase, and blocked by H89, an inhibitor of cAMP-dependent protein kinase. Additionally, we provide evidence that GLP1-stimulated activation of Erk requires an influx of calcium through L-type voltage-gated calcium channels and the activation of calcium/calmodulin-dependent protein kinase II. GLP1-stimulated activation of Erk is blocked by inhibitors of MEK, but GLP1 does not induce the activation of A-Raf, B-Raf, C-Raf, or Ras. Additionally, dominant negative forms of Ras(N17) and Rap1(N17) fail to block GLP1-stimulated activation of Erk. In conclusion, our results indicate that, in the presence of stimulatory concentrations of glucose, GLP1 stimulates the activation of Erk through a mechanism dependent on MEK but independent of both Raf and Ras. This requires 1) the activation of cAMP-dependent protein kinase, 2) an influx of extracellular Ca(2+) through L-type voltage-gated calcium channels, and 3) the activation of CaM kinase II.
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PMID:cAMP-dependent protein kinase and Ca2+ influx through L-type voltage-gated calcium channels mediate Raf-independent activation of extracellular regulated kinase in response to glucagon-like peptide-1 in pancreatic beta-cells. 1236 24


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