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)

Partially reduced oxygen species are toxic, yet activated sea urchin eggs produce H2O2, suggesting that the control of oxidant stress might be critical for early embryonic development. We show that the Ca2(+)-stimulated NADPH oxidase that generates H2O2 in the "respiratory burst" of fertilization is activated by a protein kinase, apparently to regulate the synthesis of this potentially lethal oxidant. The NADPH oxidase was separated into membrane and soluble fractions that were both required for H2O2 synthesis. The soluble fraction was further purified by anion exchange chromatography. The factor in the soluble fraction that activated the membrane-associated oxidase was demonstrated to be protein kinase C (PKC) by several criteria, including its Ca2+/phophatidylserine/diacyl-glycerol-stimulated histone kinase activity, its response to phorbol ester, its inhibition by a PKC pseudosubstrate peptide, and its replacement by purified mammalian PKC. Neither calmodulin-dependent kinase II, the catalytic subunit of cyclic AMP-dependent protein kinase, casein kinase II, nor myosin light chain kinase activated the oxidase. Although the PKC family has been ubiquitously implicated in cellular regulation, enzymes that require PKC for activation have not been identified; the respiratory burst oxidase is one such enzyme.
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PMID:A specific requirement for protein kinase C in activation of the respiratory burst oxidase of fertilization. 233 2

We have investigated the phosphorylation of the ribosomal S6 protein which may be on the pathway of mitogenic stimulation in response to oxidants. Mouse epidermal cells JB6 (clone 41) were exposed to active oxygen generated extracellularly by glucose/glucose oxidase (producing H2O2) or xanthine oxidase (producing H2O2 plus superoxide) or active oxygen produced intracellularly by the metabolism of menadione (producing mostly superoxide). All three sources of active oxygen induced rapidly a protein kinase activity which phosphorylated S6 in cellular extracts prepared in the presence of the phosphatase inhibitor beta-glycerophosphate. Maximal activity was reached within 15 min of exposure, and phosphorylation occurred specifically at serine residues. Strong activation of the protein kinase activity was also observed by diamide which selectively oxidizes SH functions. The following observations characterize the reaction: 1) Extracellular addition of catalase but not Cu,Zn-superoxide dismutase was inhibitory, implicating H2O2 rather than superoxide as the active species. 2) Exposure of JB6 cells to reagent H2O2 or H2O2 released by glucose/glucose oxidase resulted in a measurable increase in intracellular free Ca2+. 3) The intracellular Ca2+ complexer quin 2 suppressed the reaction. 4) The calmodulin antagonist trifluoperazine prevented the activation of the protein kinase. 5) Exposure of cells to Mn2+ and La3+, which stimulate calmodulin-dependent activities, potently increased the S6 kinase activity of the cell extracts. 6) Desalted extracts strictly required the addition of Mg2+ and their activity was inhibited by Mn2+. In contrast, the phosphorylation of a 95-kDa protein was strongly stimulated by Mn2+. 7) For several agonists, i.e. active oxygen, phorbol 12-myristate 13-acetate, and serum, tryptic peptide analysis yielded the same phosphopeptides, suggesting that a common S6 kinase is involved in these reactions. From these data we propose that oxidants induce an increase in intracellular free Ca2+ which activates a Ca2+/calmodulin-dependent protein kinase and, as a consequence, an S6 kinase.
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PMID:Oxidants induce phosphorylation of ribosomal protein S6. 314 21

Bivalent lectins as bridging molecules between cells or cell surface lectins as docking points are involved in mediation of cell adhesion by specific recognition of suitable glycoconjugates on an opposing surface. The initial contact formation by a lectin can lead to intracellular post-binding events which effect stable cell association even in the presence of the haptenic sugar. To delineate the participation of intracellular signaling pathways in the cascade of reactions to establish firm association, reagents with proven inhibitory capacity on certain biochemical targets provide suitable tools. Using this approach with rat thymocytes and the galactoside-binding lectin from mistletoe (Viscum album L. agglutinin, VAA) as a model, a panel of 27 inhibitors with impact on e.g. several types of kinases, tyrosine phosphatases, NO synthases, G proteins, enzymes of arachidonate and cyclic nucleotide metabolism and calmodulin was systematically tested with respect to their capacity to impair the formation of lactose-resistant cell aggregates. In addition to the recently reported effectiveness of N-ethylmaleimide, nordihydroguaiaretic acid, and trifluoperazine the agents diacylglycerol kinase inhibitor II, emodin, D609, DPI, KT5720, KT5926, MK-886, bisindolylmaleimide I, and (+/-)methoxyverapamil were able to reduce aggregate stability in the presence of the haptenic sugar. Thus, various types of kinases including p561lck tyrosine kinase, lipoxygenases, phosphatidylcholine-specific phospholipase C as well as calmodulin and Ca(2+)-currents, but not modulators of the metabolism of cyclic nucleotides, NO synthases, MAP kinases, tyrosine phosphatases and phospholipase A (preferentially group II) and C can play a role in eliciting contact stability. More than one principal signaling pathway appears to be linked to the measurable parameter, since inhibitory substances show additive properties in co-incubation assays and differentially affect two lectin-elicited cellular activities, i.e. intracellular movement of Ca(2+)-ions and H2O2-generation, which can accompany cell adhesion and aggregation. Pronounced differences in the extent of modulation of H2O2-generation in human neutrophils by the same set of substances emphasizes that general conclusions on the post-binding effects for a certain lectin in different cell types are definitely precluded. In aggregate, the approach to employ inhibitors with target selectivity intimates an involvement of protein kinases A, C, Ca2+/calmodulin-dependent protein kinase II, p56lck tyrosine kinase, leukotrienes and/or hydroxyeicosatetraenoic acids, phosphatidylcholine-specific phospholipase C and Ca(2+)-fluxes in events following initial binding of a galactoside-specific plant lectin to rat thymocytes which establish firm cell contacts.
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PMID:Dissection of the impact of various intracellular signaling pathways on stable cell aggregate formation of rat thymocytes after initial lectin-dependent cell association of using a plant lectin as model and target-selective inhibitors. 1048 33

C-Jun N-terminal kinase 1 and 2 (JNK1/2) have been shown to be transiently activated and involved in neurotoxicity. We searched for possible upstream molecules, which are responsible for the regulation of hydrogen peroxide-(H2O2) induced JNK1/2 activation and JNK1/2-mediated apoptotic-like cell death in cultured rat cortical neurons. The results showed that JNK1/2 activation (monitored by anti-diphosphorylated JNK1/2 antibody) was largely prevented by elimination of extracellular Ca2+ or blockage of NMDA-receptors (NMDA-R), and was weakly but significantly decreased by blockage of L-type voltage-gated calcium channel (L-VGCC); furthermore, JNK1/2 activation was largely prevented by inhibition of Ca2+/calmodulin-dependent protein kinase-II (CaMKII) and protein-tyrosine kinases (PTK). We also found that H2O2-induced apoptotic-like cell death was partially prevented by elimination of extracellular Ca2+, or by inhibition of NMDA-R, L-VGCC, PTK and CaMKII, respectively. The above results suggest that in H2O2-induced neurotoxicity, JNK1/2 activation is mainly mediated by NMDA-R and L-VGCC. Consequently, PTK and CaMKII are critical intermediaries in JNK1/2 activation and are mainly responsible for JNK1/2-mediated apoptotic-like cell death.
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PMID:Regulation of c-Jun N-terminal kinase activation in hydrogen peroxide induced neurotoxicity. 1470 49

Excessive excitatory amino acid (EAA) release in cerebral ischemia is a major mechanism responsible for neuronal damage and death. A substantial fraction of ischemic EAA release occurs via volume-regulated anion channels (VRACs). Hydrogen peroxide (H2O2), which is abundantly produced during ischemia and reperfusion, activates a number of protein kinases critical for VRAC functioning and has recently been reported to activate VRACs. In the present study, we explored the effects of H2O2 on volume-dependent EAA release in cultured astrocytes, measured as the release of preloaded D-[3H]aspartate. 100-1,000 microm H2O2 enhanced swelling-induced EAA release by approximately 2.5-3-fold (EC50 approximately 10 microM). The VRAC blockers ATP, phloretin, and 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB) potently inhibited both control swelling-induced and the H2O2-potentiated release, suggesting a role for VRACs. The H2O2-induced component of EAA release was attenuated by the Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester (BAPTA-AM) and completely eliminated by the calmodulin antagonists trifluoperazine and W-7 and the Ca2+/calmodulin-dependent protein kinase II (CaMKII) inhibitor KN-93. Inhibitors of tyrosine kinases, protein kinase C, and the myosin light chain kinase were ineffective in blocking the H2O2 response. H2O2 treatment of swollen astrocytes, but not swelling alone, resulted in CaMKII activation that was inhibited by KN-93, as determined by a phospho-Thr286 CaMKII antibody. These data demonstrate that H2O2 strongly up-regulates astrocytic volume-sensitive EAA release via a CaMKII-dependent mechanism and in this way may potently promote pathological EAA release and brain damage in ischemia.
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PMID:Hydrogen peroxide potentiates volume-sensitive excitatory amino acid release via a mechanism involving Ca2+/calmodulin-dependent protein kinase II. 1556 71

Angiotensin II (ANG II) promotes vascular smooth muscle cell (VSMC) growth, stimulates Ca(2+)-calmodulin (CaM)-dependent kinase II (CaMKII), and activates cytosolic Ca(2+)-dependent phospholipase A2 (cPLA2), which releases arachidonic acid (AA). ANG II also generates H2O2 and activates Akt, which have been implicated in ANG II actions in VSMC. This study was conducted to investigate the relationship of these signaling molecules to Akt activation in rat aortic VSMC. ANG II increased Akt activity, as measured by its phosphorylation at serine-473. ANG II (200 nM)-induced Akt phosphorylation was decreased by extracellular Ca2+ depletion and calcium chelator EGTA and inhibitors of CaM [N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide] and CaMKII [(2-[N-(2-hydroxyethyl)]-N-(4-me-thoxybenzenesulfonyl)]amino-N-(4-chlorocinnamyl)-N-methylbenzyl-amine)]. cPLA2 inhibitor pyrrolidine-1, antisense oligonucleotide, and retroviral small interfering RNA also attenuated ANG II-induced Akt phosphorylation. AA increased Akt phosphorylation, and AA metabolism inhibitor 5,8,11,14-eicosatetraynoic acid (ETYA) blocked ANG II- and AA-induced Akt phosphorylation (199.03 +/- 27.91% with ANG II and 110.18 +/- 22.40% with ETYA + ANG II; 405.00 +/- 86.22% with AA and 153.97 +/- 63.26% with ETYA + AA). Inhibitors of lipoxygenase (cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate) and cytochrome P-450 (ketoconazole and 17-octadecynoic acid), but not cyclooxygenase (indomethacin), attenuated ANG II- and AA-induced Akt phosphorylation. Furthermore, 5(S)-, 12(S)-, 15(S)-, and 20-hydroxyeicosatetraenoic acids and 5,6-, 11,12-, and 14,15-epoxyeicosatrienoic acids increased Akt phosphorylation. Catalase inhibited ANG II-increased H2O2 production but not Akt phosphorylation. Oleic acid, which also increased H2O2 production, did not cause Akt phosphorylation. These data suggest that ANG II-induced Akt activation in VSMC is mediated by AA metabolites, most likely generated via lipoxygenase and cytochrome P-450 consequent to AA released by CaMKII-activated cPLA2 and independent of H2O2 production.
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PMID:Angiotensin II-induced Akt activation is mediated by metabolites of arachidonic acid generated by CaMKII-stimulated Ca2(+)-dependent phospholipase A2. 1563 21

The understanding of endothelial cell responses to oxidative stress may provide insights into aging mechanisms and into the pathogenesis of numerous cardiovascular diseases. In this study, we examined the regulation and the functional role of cyclin D1, a crucial player in cell proliferation and survival. On H2O2 treatment, endothelial cells showed a rapid down-modulation of cyclin D1. Other D-cyclins were similarly regulated, and this decrease was also observed after exposure to other oxidative stress-inducing stimuli, namely 1,3-bis (2 chloroethyl)-1 nitrosourea treatment and ischemia. H2O2 treatment induced cyclin D1 ubiquitination followed by proteasome degradation. Phospholipase C inhibition prevented cyclin D1 degradation, and its activation triggered cyclin D1 down-modulation in the absence of oxidative stress. Activated phospholipase C generates inositol-1,4,5-trisphosphate (IP3) and Ca2+ release from internal stores. We found that both IP3-receptor inhibition and intracellular Ca2+ chelation prevented cyclin D1 degradation induced by oxidative stress. Furthermore, Ca2+ increase was transduced by Ca2+/calmodulin-dependent protein kinase (CaMK). In fact, H2O2 stimulated CaMK activity, CaMK inhibitors prevented H2O2-induced cyclin D1 down-modulation, and CaMK overexpression induced cyclin D1 degradation. Finally, overriding of cyclin D1 down-modulation via its forced overexpression or via CaMK inhibition increased cell sensitivity to H2O2-induced apoptotic cell death. Thus, cyclin D1 degradation enhances endothelial cell survival on oxidative stress.
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PMID:Cyclin D1 degradation enhances endothelial cell survival upon oxidative stress. 1660 4

The present study was undertaken to evaluate the implication of delta-opioid receptor function in neurogenesis and neuroprotection. We found that the stimulation of delta-opioid receptors by the selective delta-opioid receptor agonist SNC80 [(+)-4-[(alphaR)-alpha-((2S,5R)-4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl]-N,N-diethylbenzamide] (10 nm) promoted neural differentiation from multipotent neural stem cells obtained from embryonic C3H mouse forebrains. In contrast, either a selective micro-opioid receptor agonist, [D-Ala2, N-Me-Phe4, Gly5-ol]-enkephalin (DAMGO), or a specific kappa-opioid receptor agonist, (-)-trans-(1S,2S)-U-50488 hydrochloride (U50,488H), had no such effect. In addition to neural differentiation, the increase in cleaved caspase 3-like immunoreactivity induced by H2O2 (3 microm) was suppressed by treatment with SNC80 in cortical neuron/glia co-cultures. These effects of SNC80 were abolished by a Trk-dependent tyrosine kinase inhibitor: (8R*,9S*,11S*)-(-)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8,11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo(a,g)cycloocta(cde)trinden-1-one (K-252a). The SNC80-induced neural differentiation was also inhibited by treatment with the protein kinase C (PKC) inhibitor, phosphatidylinositol 3-kinase (PI3K) inhibitor, mitogen-activated protein kinase kinase (MEK) inhibitor or Ca2+/calmodulin-dependent protein kinase II (CaMKII) inhibitor. These findings raise the possibility that delta-opioid receptors play a crucial role in neurogenesis and neuroprotection, mainly through the activation of Trk-dependent tyrosine kinase, which could be linked to PI3K, PKC, CaMKII and MEK.
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PMID:Role of delta-opioid receptor function in neurogenesis and neuroprotection. 1669 56

In response to metabolic stress, GLUT4, the most abundant glucose transporter, translocates from intracellular vesicles to the plasma membrane. This appears to play an important role in protecting cardiac myocytes from ischemic injury. To investigate the precise mechanisms of GLUT4 translocation in cardiomyocytes, we have established a method for quantifying the relative proportion of sarcolemmal GLUT4 to total GLUT4 in these cells. Stimulation with H2O2 resulted in a concentration-dependent increase in GLUT4 translocation, which peaked at 15 min after stimulation. The dominant-negative form (DN) of AMP-activated protein kinase (AMPK) alpha2 inhibited the H2O2-induced translocation of GLUT4. We further examined the role of two known AMPK kinases (AMPKKs), calmodulin-dependent protein kinase kinase (CaMKK)beta and LKB1. The DN of CaMKKbeta or LKB1 alone inhibited H2O2-induced GLUT4 translocation only partially compared to the inhibition produced by the DN of AMPKalpha2. However, the combination of DN-LKB1 and DN-CaMKKbeta inhibited translocation to an extent similar to with DN-AMPKalpha2. Stimulation with H2O2 also activated Akt and the inhibition of PI3-K/Akt prevented GLUT4 translocation to the same extent as with AMPK inhibition. When the DN of AMPKalpha2 was applied with DN-PI3-K, there was a complete reduction in the GLUT4 membrane level similar to that seen at the 0 time-point. These results demonstrate that AMPK and PI3-K/Akt have an additive effect on oxidative stress-mediated GLUT4 translocation.
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PMID:Oxidative stress induces GLUT4 translocation by activation of PI3-K/Akt and dual AMPK kinase in cardiac myocytes. 1816 80

Ca2+/calmodulin (CaM)-dependent protein kinase II (CaMKII) plays a critical role in neuronal signal transduction and synaptic plasticity. Here, we showed that this kinase was very susceptible to oxidative modulation. Treatment of mouse brain synaptosomes with H2O2, diamide, and sodium nitroprusside caused aggregation of CaMKII through formation of disulfide and non-disulfide linkages, and partial inhibition of the kinase activity. These CaMKII aggregates were found to associate with the post synaptic density. However, treatment of purified CaMKII with these oxidants did not replicate those effects observed in the synaptosomes. Using two previously identified potential mediators of oxidants in the brain, glutathione disulfide S-monoxide (GS-DSMO) and glutathione disulfide S-dioxide (GS-DSDO), we showed that they oxidized and inhibited CaMKII in a manner partly related to those of the oxidant-treated synaptosomes as well as the ischemia-elicited oxidative stress in the acutely prepared hippocampal slices. Interestingly, the autophosphorylated and activated CaMKII was relatively refractory to GS-DSMO- and GS-DSDO-mediated aggregation. Short term ischemia (10 min) caused a depression of basal synaptic response of the hippocampal slices, and re-oxygenation (after 10 min) reversed the depression. However, oxidation of CaMKII remained at above the pre-ischemic level throughout the treatment. Oxidation of CaMKII also prevented full recovery of CaMKII autophosphorylation after re-oxygenation. Subsequently, the high frequency stimulation-mediated synaptic potentiation in the hippocampal CA1 region was significantly reduced compared with the control without ischemia. Thus, ischemia-evoked oxidation of CaMKII, probably via the action of glutathione disulfide S-oxides or their analogues, may be involved in the suppression of synaptic plasticity.
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PMID:Ischemia-elicited oxidative modulation of Ca2+/calmodulin-dependent protein kinase II. 1817 65


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