Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.17 (CaMKII)
 4,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Posttranslational modifications of synapsin I, a major phosphoprotein in synaptic terminals, were studied by mass spectrometry. In addition to a well known phosphorylation site by calmodulin-dependent protein kinase II (CaM kinase II), a hitherto unrecognized site (Ser553) was found phosphorylated in vivo. The phosphorylation site is immediately followed by a proline, suggesting that the protein is an in vivo substrate of so-called proline-directed protein kinase(s). To identify the kinase involved, three proline-directed protein kinases expressed highly in the brain, i.e. mitogen-activated protein (MAP) kinase, Cdk5-p23, and glycogen synthase kinase 3beta, were tested for the in vitro phosphorylation of synapsin I. Only MAP kinase and Cdk5-p23 phosphorylated synapsin I stoichiometrically. The phosphorylation sites were determined to be Ser551 and Ser553 with Cdk5-p23, and Ser62, Ser67, and Ser551 with MAP kinase. Upon phosphorylation with MAP kinase, synapsin I showed reduced F-actin bundling activity, while no significant effect on the interaction was observed with the protein phosphorylated with Cdk5-p23. These results raise the possibility that the so-called proline-directed protein kinases together with CaM kinase II and cAMP-dependent protein kinase play an important role in the regulation of synapsin I function.
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PMID:Site-specific phosphorylation of synapsin I by mitogen-activated protein kinase and Cdk5 and its effects on physiological functions. 870 79

Many of the extracellular signals such as neurotransmitters and hormones regulate the intracellular concentration of second messengers such as cAMP, cGMP, and calcium ion (Ca2+), diacylglycerol and IP3. Accumulating evidence indicates that protein phosphorylation-dephosphorylation is an important mechanism by which second messengers act to regulate a variety of cellular processes. Ca2+/calmodulin-dependent protein kinase II, cAMP-dependent protein kinase and protein kinase C are three major classes of protein kinases in the central nervous system. In an attempt to elucidate the physiological roles of the protein kinases, I have been studying the substrate proteins and functional significance of the enzymes and phosphorylated proteins. For these purposes, I investigated the phosphorylation-dephosphorylation of cytoskeletal proteins such as microtubule-associated protein 2 and tau, which are involved in the assembly-disassembly of microtubules and the production of abnormally phosphorylated forms of tau in neurofibrillary tangles in Alzheimer's disease brain. As the natural consequence, studying the protein phosphatases is significant for elucidating the switch-off mechanism of protein phosphorylation. Thus, I have been investigating the functional significance of protein phosphorylation-dephosphorylation for the elucidation of signal transduction in the brain, which is widely involved in the regulation of brain functions.
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PMID:[Molecular and pharmacological studies on signal transduction in the brain]. 872 Feb 94

Inhibition of type III adenylyl cyclase (III-AC) by intracellular Ca2+ in vivo provides a mechanism for attenuation of hormone-stimulated cAMP signals in olfactory epithelium, heart, and other tissues (Wayman, G. A., Impey, S., and Storm, D. R. (1995) J. Biol. Chem. 270, 21480-21486). Although the mechanism for Ca2+ inhibition of III-AC in vivo has not been defined, inhibition is not mediated by Gi, cAMP-dependent protein kinase, or protein kinase C. However, Ca2+ inhibition of III-AC is antagonized by KN-62, a CaM-dependent kinase inhibitor. In addition, constitutively activated CaM kinase II inhibits the enzyme. These data suggest that CaM kinase II regulates the activity of III-AC by direct phosphorylation or by an indirect mechanism involving phosphorylation of a protein that inhibits III-AC. Here we report that III-AC is phosphorylated in vivo when intracellular Ca2+ is increased and that phosphorylation is prevented by CaM-dependent kinase inhibitors. Site-directed mutagenesis of a CaM kinase II consensus site (Ser-1076 to Ala-1076) in III-AC greatly reduced Ca2+-stimulated phosphorylation and inhibition of III-AC in vivo. These data support the hypothesis that Ca2+ inhibition of III-AC is due to direct phosphorylation of the enzyme by CaM kinase II in vivo.
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PMID:Phosphorylation and inhibition of type III adenylyl cyclase by calmodulin-dependent protein kinase II in vivo. 879 67

ATP evoked whole-cell potassium currents in hippocampal neurons. The second application of ATP to the same cell potentiated the current amplitude to around 140% and the current potentiation was maintained by further applications. A calmodulin inhibitor, W-7, or a selective Ca2+/calmodulin-dependent protein kinase II (CaMKII) inhibitor, KN-62, inhibited the current potentiation, although a selective protein kinase C inhibitor, GF109203X, or a selective cAMP-dependent protein kinase inhibitor, H-89, had no effect. In addition, ATP enhanced intracellular free Ca2+ concentration, which may activate CaMKII, but this enhancement was blocked by repetitive applications. These results provide an indication that CaMKII may be involved in the current potentiation by repetitive applications of ATP.
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PMID:Repetitive applications of ATP potentiate potassium current by Ca2+/calmodulin kinase in cultured rat hippocampal neurons. 883 7

Relaxin, a reproductive hormone of the insulin-like growth factor family, increases heart rate in experimental animals but its other actions on cardiac function and cellular mechanisms responsible for the positive chronotrophic effect remain unknown. We have studied the actions of human recombinant gene-2 relaxin on the release of atrial natriuretic peptide (ANP) and cardiac function (heart rate, contractile force, perfusion pressure) as well as the underlying signal transduction mechanisms by using the isolated perfused spontaneously beating rat heart preparation. The administration of relaxin into the perfusion fluid at concentrations of 1.5, 3 or 10 nM for 30 min caused a dose-dependent sustained increase in heart rate, while contractile force and perfusion pressure remained unchanged. In addition, infusion of relaxin at a concentration of 10 nM into the perfusate produced a gradual 1.5-fold increase in immunoreactive ANP (IR-ANP) secretion (from 456 +/- 76 to 701 +/- 124 pg/ml, F = 4.5, P < 0.001). The ANP secretory and chronotrophic effects of relaxin appear to involve the activation of protein kinase C, since administration of a protein kinase C inhibitor staurosporine at a concentration of 30 nM completely blocked the effect of relaxin (10 nM) on IR-ANP secretion (P < 0.001) and heart rate (P < 0.001). A cAMP-dependent protein kinase inhibitor, H-89 (100 nM), also substantially reduced the ANP secretory effect of relaxin and attenuated the increase in heart rate during the sustained phase of the relaxin infusion (P < 0.001). KN-62 (3 microM), a Ca2+/calmodulin-dependent protein kinase inhibitor, decreased the positive chronotrophic effect of relaxin (P < 0.001) but did not influence significantly the effect of relaxin on IR-ANP release in isolated perfused rat heart preparation. These results provide the first evidence that relaxin stimulates the secretion of ANP from isolated perfused rat hearts. Our results also suggest that relaxin modulates ANP secretion by activation of protein kinase C and cAMP-dependent protein kinase pathways.
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PMID:Relaxin stimulates atrial natriuretic peptide secretion in perfused rat heart. 888 68

In the adult myocardium the Ca2+ uptake and release functions of the sarcoplasmic reticulum (SR) are known to be regulated by a membrane-associated Ca2+-calmodulin-dependent protein kinase (CaM kinase) which phosphorylates the Ca2+-pumping ATPase (Ca2+ pump), Ca2+ release channel (ryanodine receptor) and the Ca2+ pump-regulatory protein, phospholamban. The role of CaM kinase during development, however, has not been examined previously. The present study investigated the ontogenetic expression of SR-associated CaM kinase in the rabbit myocardium as well as development-related changes in CaM kinase-mediated phosphorylation of the SR proteins (Ca2+ pump, Ca2+ release channel and phospholamban) involved in transmembrane Ca2+ cycling. For these experiments, cardiac muscle homogenate and SR-enriched membrane fraction derived from fetal (21- and 28-days gestation), newborn (2 days postnatal) and adult New Zealand White rabbits were used. Western immunoblotting analysis detected the presence of phospholamban, Ca2+ pump and Ca2+ release channel in homogenate and SR at all ages tested. The amount of these proteins in the SR increased substantially during fetal and postnatal development. Phosphorylation studies revealed the presence of CaM kinase-dependent phosphorylation of the Ca2+ pump, Ca2+ release channel and phospholamban as early as 21-days gestation. This phosphorylation could be elicited with the addition of only Ca2+ and calmodulin indicating the presence of a SR-associated CaM kinase as early as 21-days gestation. This was confirmed using a delta-CaM kinase II-specific antibody. Phosphorylation per unit amount of each substrate was greater in the fetus and newborn compared to adult. Phosphorylation of phospholamban could be elicited by exogenous cAMP-dependent protein kinase (PKA) at all developmental stages studied. Activation of SR CaM kinase with Ca2+ and calmodulin, or induction of phospholamban phosphorylation by exogenous PKA, resulted in stimulation of the Ca2+ uptake activity of SR in fetal, newborn and adult heart. These results demonstrate early ontogenetic expression of the Ca2+ cycling proteins and CaM kinase in the SR and the concurrent development of phosphorylation-dependent regulation of SR Ca2+ cycling.
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PMID:Ontogeny of sarcoplasmic reticulum protein phosphorylation by Ca2+--calmodulin-dependent protein kinase. 904 54

Phospholamban (PLB), the regulator of the cardiac sarcoplasmic reticulum (SR) Ca2+ pump is specifically phosphorylated at Ser16 and Thr17 by cAMP-dependent protein kinase (PKA) and Ca2+/calmodulin-dependent protein kinase (CaMK), respectively. The regulation of this dual-site phosphorylation of amino acid residues in direct proximity is only poorly understood. In order to study the site-specific phosphorylation of PLB, we used a synthetic peptide (PLB-24) corresponding to the cytosolic part of the PLB monomer with the phosphorylation sites as a model substrate. PLB-24 possesses substrate properties as the native PLB as demonstrated by phosphorylation with exogenous, purified PKA, cGMP-dependent protein kinase (PKG) and a type II CaMK (CaMKII). In isolated vesicles of cardiac SR there was a rapid phosphorylation of the peptide by the endogenous PKA (SR-PKA) and CaMK (SR-CaMK), but not under conditions that activate PKG. Both SR-PKA and SR-CaMK incorporated the same amount of 32P into PLB-24, 0.60 +/- 0.01 nmol 32P/mg SR protein and 0.61 +/- 0.03 nmol 32P/mg SR protein, respectively. Phosphorylation by SR-PKA was abolished by the specific PKA inhibitor (IC50 = 0.2 microM), whereas SR-CaMK phosphorylation was inhibited by calmidazolium (IC50 = 1.6 microM) and a CaMKII-specific inhibitor peptide (IC50 = 2.5 microM). Phosphorylation by SR-PKA was exclusively at Ser, whereas SR-CaMK phosphorylated only Thr. After simultaneous activation of both SR-kinases 32P incorporation into PLB-24 was additive and occurred at Ser as well as at Thr. Sequential activation of SR-PKA and SR-CaMK also caused the additive phosphorylation of PLB-24 independently of which kinase was activated first. Thus, at the monomeric level of PLB the respective phosphorylation site appears to be accessible to its related SR protein kinase in vitro even when the adjacent site is phosphorylated.
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PMID:Site-specific phosphorylation of a phospholamban peptide by cyclic nucleotide- and Ca2+/calmodulin-dependent protein kinases of cardiac sarcoplasmic reticulum. 920 42

The present work was undertaken to determine the action of methylmalonic acid (MMA), a metabolite, which accumulates in high amounts in methylmalonic acidemia, on the endogenous phosphorylating system associated with the cytoskeletal fraction proteins of cerebral cortex of young rats. We demonstrated that pre-treatment of cerebral cortex slices of young rats with 2.5 mM buffered methylmalonic acid (MMA) is effective in decreasing in vitro incorporation of [32P]ATP into neurofilament subunits (NF-M and NF-L) and alpha- and beta-tubulins. Based on the fact that this system contains cAMP-dependent protein kinase (PKA), Ca2+/calmodulin-dependent protein kinase II (CaMKII) and protein phosphatase 1 (PP1), we first tested the effect of MMA on the kinase activities by using the specific activators cAMP and Ca2+/calmodulin or the inhibitors PKAI or KN-93 for PKA and CaMKII, respectively. We observed that MMA totally inhibited the stimulatory effect of cAMP and interfered with the inhibitory effect of PKAI. In addition, the metabolite partially prevented the stimulatory effect of Ca2+/calmodulin and interfered with the effect of KN-93. Furthermore, in vitro dephosphorylation of neurofilament subunits and tubulins was totally inhibited in brain slices pre-treated with MMA. Taken together, these results suggest that MMA, at the same concentrations found in tissues of methylmalonic acidemic children, inhibits the in vitro activities of PKA, CaMKII and PP1 associated with the cytoskeletal fraction of the cerebral cortex of rats, a fact that may be involved with the pathogenesis of the neurological dysfunction characteristic of methylmalonic acidemia.
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PMID:Methylmalonic acid reduces the in vitro phosphorylation of cytoskeletal proteins in the cerebral cortex of rats. 929 63

We have demonstrated that treatment with 200 nM okadaic acid (OA) for 1 h followed by a 15-min heat shock (HS) at 45 degrees C (termed OA-->HS treatment) leads to a rapid transactivation of grp78, the gene for the 78-kDa glucose-regulated protein, in 9L rat brain tumor cells. The level of Grp78 mRNA rose 15-fold in 60 min after the combined treatment. Nuclear extracts from cells subjected to OA-->HS treatment, compared to those of treatment with OA or HS alone, exhibited an increased binding activity toward an oligonucleotide probe containing the cAMP-responsive element-like (CRE-like, TGACGTGA) regulatory element in electrophoretic mobility shift assays (EMSA). The binding resulted in the formation of two protein-EMSA probe complexes exhibiting different association and dissociation rates in kinetic studies. The protein factors in the upper band (complex I) and lower band (complex II) were identified as the activating transcription factor-2 (ATF-2) and the CRE binding factor 1 (CREB-1), respectively, by antibody interference assays. In addition, the identity of CREB-1 was confirmed by supershift analysis. The binding activity, as well as the transactivation of the grp78 gene, can be abolished by a 1-h treatment with the cAMP-dependent protein kinase (PKA) inhibitor but not with protein kinase C or Ca2+/calmodulin-dependent protein kinase II inhibitors. Accumulation of steady-state level of ATF-2 was observed and was also modulated by treatment with H-89, a PKA inhibitor. From these results, we conclude that the CRE-like element plays an important role in the rapid transactivation of the grp78 gene and that the PKA signaling pathway is involved. In addition, PKA-mediated transcriptional regulation of grp78 in OA-->HS treatment is through regulation of protein phosphorylation as well as de novo synthesis of ATF-2.
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PMID:Rapid induction of the Grp78 gene by cooperative actions of okadaic acid and heat-shock in 9L rat brain tumor cells--involvement of a cAMP responsive element-like promoter sequence and a protein kinase A signaling pathway. 931 Mar 69

cAMP and Ca2+ acted together with the acute phase cytokine interleukin-1beta (IL-1beta) to inhibit hepatocyte DNA replication. At sub-basal activity of cAMP-dependent protein kinase (PKA), neither IL-1beta nor the Ca2+-elevating hormone vasopressin affected hepatocyte proliferation. Basal level of PKA activity permitted IL-1beta action. Increased PKA activity also permitted vasopressin action and sensitized further towards IL-1beta, which acted at 10-50 pM concentrations. Vasopressin acted via Ca2+/calmodulin-dependent protein kinase II (CaMKII), and its action was mimicked by the serine/threonine phosphatase inhibitor microcystin, which activates CaMKII. Inhibitors (KN93 and KT5926) of CaMKII selectively counteracted the effects of vasopressin and microcystin on hepatocyte proliferation at concentrations similar to those required to inhibit CaMKII in vitro. Two-dimensional gel electrophoresis of 32P-prelabeled hepatocytes revealed a common set of proteins phosphorylated in response to vasopressin and microcystin. Their phosphorylation was counteracted by CaMKII inhibitor (KT5926). Phosphorylation of the CaMKII substrate phenylalanine hydroxylase (PAH; EC 1.14.16.1) was used as an endogenous marker of CaMKII activation. It was found that treatment of the cells with vasopressin or microcystin increased the phosphorylation of PAH, and that the vasopressin-induced PAH phosphorylation was inhibited by KT5926. In conclusion, the Ca2+-elevating hormone vasopressin potentiated the antiproliferative effects of cAMP and IL-1beta through CaMKII activation.
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PMID:Synergistic antiproliferative actions of cyclic adenosine 3',5'-monophosphate, interleukin-1beta, and activators of Ca2+/calmodulin-dependent protein kinase in primary hepatocytes. 932 53


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