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 investigated specific signaling events initiated after T cell triggering through the costimulatory surface receptors CD2 and CD28 as compared with activation via the Ag receptor (TCR/CD3). We therefore followed the phosphorylation of stathmin, a ubiquitous cytoplasmic phosphoprotein proposed as a general relay integrating diverse intracellular signaling pathways through the combinatorial phosphorylation of serines 16, 25, 38, and 63, the likely physiologic substrates for Ca2+/calmodulin (CaM)-dependent kinases, mitogen-activated protein (MAP) kinase, cyclin-dependent kinases (cdks), and protein kinase A, respectively. We addressed the specific protein kinase systems involved in the CD2 pathway of T cell activation through the analysis of stathmin phosphorylation patterns in exponentially growing Jurkat T cells, as revealed by phosphopeptide mapping. Stimulation via CD2 activated multiple signal transduction pathways, resulting in phosphorylation of distinct sites of stathmin, the combination of which only partially overlaps the CD3- and CD28-induced patterns. The partial redundancy of the three T cell activation pathways was evidenced by the phosphorylation of Ser25 and Ser38, substrates of MAP kinases and of the cdk family kinase(s), respectively. Conversely, the phosphorylation of Ser16 of stathmin was observed in response to both CD2 and CD28 triggering, but not CD3 triggering, with a kinetics compatible with the lasting activation of CaM kinase II in response to CD2 triggering. In vitro, Ser16 of recombinant human stathmin was phosphorylated also by purified CaM kinase II, and in vivo, CaM kinase II activity was indeed stimulated in CD2-triggered Jurkat cells. Altogether, our results favor an association of CaM kinase II activity with costimulatory signals of T lymphocyte activation and phosphorylation of stathmin on Ser16.
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PMID:Serine 16 of stathmin as a cytosolic target for Ca2+/calmodulin-dependent kinase II after CD2 triggering of human T lymphocytes. 968 69

The present study was undertaken to reveal underlying mechanisms of apoptosis in neurons using clonal neuronal cells, ML-DmBG2-c2, derived from Drosophila larval central nervous system 1-(5-Isoquinolinesulfonyl)-2-methylpiperazine (H-7), a protein kinase inhibitor, induced cell death with typical features of apoptosis such as internucleosomal DNA fragmentation, nuclear condensation and apoptotic bodies in the cells. Though H-7 is known to inhibit cAMP-dependent protein kinase (PKA), protein kinase C (PKC), cGMP-dependent protein kinase (PKG), myosin light chain kinase (MLCK), and casein kinase I (CKI), specific inhibitors for these kinases such as H-89, calphostin C, ML-9, or CKI-7 did not induce apoptosis in the cells. Other kinases such as tyrosine kinase. PI3-kinase and Ca2+/CaM kinase II so far examined in the present study were interpreted not to be involved in the apoptotic cascade. Therefore, it is concluded that an H-7-sensitive substance(s) other than these kinases is responsible for the apoptosis in the neuronal cells. Caspase inhibitors prevented apoptosis in the cells treated with H-7. These results suggest that caspase(s) is involved downstream of the H-7-sensitive point in the cascade of the apoptosis.
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PMID:H-7-induced apoptosis in the cells of a Drosophila neuronal cell line through affecting unidentified H-7-sensitive substance(s). 970 Jul 17

The paired helical filaments (PHFs) found in Alzheimer's disease (AD) brains are composed primarily of the microtubule-associated protein tau. PHF-tau is in a hyperphosphorylated state and is unable to promote microtubule assembly. We investigated whether the inhibition of tau binding to microtubules is increased when tau is phosphorylated by different kinases in combination with GSK-3. We found that when tau was first phosphorylated by A-kinase, C-kinase, cdk5, or CaM kinase II and then by GSK-3, its binding to microtubules was inhibited by 45, 61, 78, and 79%, respectively. Further, the kinase combinations cdk5/GSK-3 and CaM kinase II/GSK-3 rapidly phosphorylated the sites Thr 231 and Ser 235. When these sites were individually replaced by Ala and the phosphorylation experiments repeated, tau binding to microtubules was inhibited by 54 and 71%, respectively. By comparison, when Ser 262 was replaced by Ala, tau binding to microtubules was inhibited by only 8% after phosphorylation by CaM kinase II. From these observations we estimate that the phosphorylation of Thr 231, Ser 235, and Ser 262 contributes approximately 26, approximately 9, and approximately 33%, respectively, of the overall inhibition of tau binding to microtubules. Together, our results indicate that the binding of tau to microtubules is controlled by the phosphorylation of several sites, among which are Thr 231, Ser 235, and Ser 262.
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PMID:Phosphorylation of tau at both Thr 231 and Ser 262 is required for maximal inhibition of its binding to microtubules. 973 71

Four NPY receptor subtypes have been cloned, and shown to be coupled to both Ca2+ and cAMP. However, very little is known about the downstream elements mediating NPY actions. It has recently been demonstrated in our laboratory that intrahypothalamic (i.h.t.) administration of NPY induces hypothalamic CaM kinase activity, cyclic AMP response element binding protein (CREB) phosphorylation and cyclic AMP response element (CRE) binding activity in rat hypothalamic nuclear proteins. In the present study, we have investigated whether these changes in CRE binding transcriptional factors activated by NPY results in gene regulation using a human neuroblastoma cell line (SK-N-BE2). This cell line which expresses the Y2 subtype of NPY receptors was transfected with a fusion gene containing 1.305 kb of human CRF 5' flanking region with a perfect palindromic CRE site linked to firefly luciferase gene. NPY treatment increased CaM kinase II activity, CREB phosphorylation and CRE binding in these cells. In transfected cells, luciferase activity was also increased by NPY (1.8-4-fold) within 4 h of treatment. Moreover, forskolin (7-30-fold), which stimulates cAMP production, and thapsigargin (6-8-fold), which mobilizes intracellular calcium, also increased luciferase activity within 4 h of treatment. PMA (phorbol-12-myristate-13-acetate), an activator of protein kinase-C, induced luciferase activity by 1.8-fold. NPY augmented forskolin-stimulated luciferase activity from 11- to 15-fold, but had no significant effect on thapsigargin-induced luciferase activity. These findings suggest that activation of protein kinase A (PKA) or CaM kinase leads to the induction of fusion gene. NPY treatment upregulated fusion gene expression through Ca2+ pathway in SK-N-BE2 cell line. Pretreatment with CREB antisense, but not the sense oligodeoxynucleotides, inhibited forskolin-, thapsigargin- and NPY-stimulated luciferase activity. However, CREB sense or antisense oligodeoxynucleotide treatment had no effect on PMA-stimulated luciferase activity. Furthermore, NPY induced CRE binding activity and the expression of CRE containing Y1 receptor gene in SK-N-MC cell line. These findings suggest that NPY can upregulate CRE containing reporter gene including Y1 receptor gene and NPY-induced reporter gene regulation in SK-N-BE2 cells is mediated by intracellular Ca2+ and CREB protein.
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PMID:NPY upregulates genes containing cyclic AMP response element in human neuroblastoma cell lines bearing Y1 and Y2 receptors: involvement of CREB. 980 24

Transfected Chinese hamster ovary cells expressing the rat neurotensin receptor (CHO-NTR cells) were used to study the 'Ca2+ stores depletion-Ca2+ entry' coupling which follows stimulation with neurotensin and liberation of inositol 1,4,5-trisphosphate. This coupling could be dissociated in time: the stores were emptied by stimulation with neurotensin in the absence of extracellular Ca2+; thereafter, readmission of extracellular Ca2+ produced a transient entry of Ca2+ that was progressively restored in the endoplasmic reticulum. We showed previously that the rise of [Ca2+]i during Ca2+ stores depletion controls the subsequent entry of Ca2+ and that unknown protein kinases and phosphatases may also be involved in this coupling. Here we show that: 1. W-7 (25 microM), KN-62 (10 microM) and a myristoylated autocamtide-2 related inhibitory peptide (20 microM), three inhibitors of the calcium-calmodulin-dependent protein kinase II (CaM kinase II) inhibit the entry of Ca2+ induced by emptying the stores of Ca2+ with neurotensin and thapsigargin. 2. Ca2+ stores depletion-Ca2+ entry coupling is also greatly diminished by 10 microM ONO-RS-082, an inhibitor of the phospholipase A2 (PLA2). 3. Arachidonic acid (5-100 microM) produces an entry of Ca2+; the same result is obtained by use of 5, 8, 11, 14-eicosatetraynoic acid, a non-metabolizable analog of arachidonic acid. 4. NTR-CHO cells are labeled with [3H] arachidonic acid for 24 h (progressively incorporated in membrane phospholipids). Upon neurotensin (1 nM) and thapsigargin (1 microM) stimulation, these cells produce a release of arachidonic acid which lasts for as long as the stores are empty and stops when they are reloaded with Ca2+. This production of arachidonic acid is significantly diminished by suppressing the [Ca2+]i transient during stores depletion (with cell permeant EGTA), by the PLA2 inhibitor ONO-RS-082 (10 microM) and by the CaM kinase II inhibitor KN-62 (10 microM). 5. The rise of [Ca2+]i by itself (induced by flash photolysis of nitrophenyl-EGTA), i.e. without depletion of the stores, is not sufficient to trigger an entry of Ca2+. 6. The reloading process of Ca2+ into the endoplasmic reticulum is inhibited by 10 microM chelerythrine, 100 nM GF 109203X, two inhibitors of protein kinases C (PKC) or by their downregulation by a prolonged treatment of the cells with 1 microM phorbol-12, 13-dibutyrate. We therefore suggest the involvement of CaM kinase II and PLA2 in the 'Ca2+ stores depletion-Ca2+ entry' coupling in these transfected CHO cells.
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PMID:Ca2+ entry in CHO cells, after Ca2+ stores depletion, is mediated by arachidonic acid. 988 83

The aim of the present study was to investigate whether the activities of the renal basolateral organic anion transporter (PAH transporter) and the sodium-dependent dicarboxylate transporter are modulated by the calcium/calmodulin-dependent multifunctional protein kinase II (CaM kinase II). The studies were performed on isolated S2 segments of proximal tubules microdissected from rabbit kidneys without the use of enzymatic agents. 3H-PAH was used as marker substance of the anion transporter, and 14C-glutarate as a marker of the sodium/dicarboxylate cotransporter. Because the tubules were not perfused, and hence were collapsed, the tubular uptake of the marker substances reflects transport across the basolateral cell membrane. To obtain uptake rates most closely related to initial transport rates, 30 s tubular uptake measurements were performed. The results show that a selective inhibitor of CaM kinase II, KN93, inhibited tubular PAH uptake. The smallest effective dose was 10(-7) M. An inactive analogue of KN93, KN92, was without effect, even at the high concentration of 10(-5) M. In contrast to PAH transport, tubular 14C-glutarate uptake was not affected by KN93 (10(-5) M). PAH transport was also inhibited after elevation of intracellular Ca2+ by the Ca(2+)-ionophore A 23187 and by the polycationic antibiotic neomycin, but not by the intracellular Ca2+ modulators thapsigargin and ryanodine. The effect of the Ca(2+)-ionophore could be abolished by KN93, but not by Rp-cAMPs, an inhibitor of protein kinase A, indicating that this event was mediated by CaM kinase II, but not by PKA. The results provide the first evidence that, in addition to the protein kinases A and C (previous studies from this lab), CaM kinase II has a role in the regulation of the renal basolateral PAH transporter, whereas the renal basolateral dicarboxylate transporter does not depend on CaM kinase II activity.
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PMID:Role of the calcium/calmodulin-dependent protein kinase II in the regulation of the renal basolateral PAH and dicarboxylate transporters. 1002 89

A discovery approach based on an intramolecular inhibitory mechanism was applied to a prototype calmodulin (CaM)-regulated protein kinase in order to demonstrate a proof-of-principle for the development of selective inhibitors. The overall approach used functional genomics analysis of myosin light chain kinase (MLCK) to identify short autoinhibitory sequences that lack CaM recognition activity, followed by recursive combinatorial peptide library production and comparative activity screens. Peptide 18 (Arg-Lys-Lys-Tyr-Lys-Tyr-Arg-Arg-Lys-NH2), one of several selective inhibitors discovered, has an IC50 = 50 nM for MLCK, inhibits CaM kinase II only at 4000-fold higher concentrations, and does not inhibit cyclic AMP-dependent protein kinase. Analogues of peptide 18 containing conformationally constrained cis-4-aminocyclohexanecarboxylic acid retained affinity and selectivity. The inhibitors add to the armamentarium available for the deconvolution of complex signal transduction pathways and their relationship to homeostasis and disease, and the approach is potentially applicable to enzymes in which the catalytic and regulatory domains are found within the same open reading frame of a cDNA.
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PMID:Identification of novel classes of protein kinase inhibitors using combinatorial peptide chemistry based on functional genomics knowledge. 1007 88

Increasing evidence supports a physiological role of Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) in the secretion of insulin from the pancreatic beta-cell, but the precise sites of action are not known. A role of this enzyme in neuroexocytosis is implicated by its phosphorylation of a vesicle-associated protein, synapsin I. Because of emerging similarities to the neuron with respect to exocytotic mechanisms, the expression and phosphorylation of synapsin I in the beta-cell have been studied. Synapsin I expression in clonal mouse beta-cells (betaTC3) and primary rat islet beta-cells was initially confirmed by immunoblot analysis. By immunoprecipitation, in situ phosphorylation of synapsin I was induced in permeabilized betaTC3 cells within a Ca2+ concentration range shown to activate endogenous CaM kinase II under identical conditions. Proteolytic digests of these immunoprecipitates revealed that calcium primarily induced the increased phosphorylation of sites identified as CaM kinase II-specific and distinct from protein kinase A-specific sites. Immunofluorescence and immunogold electron microscopy verified synapsin I expression in betaTC3 cells and pancreatic slices but demonstrated little if any colocalization of synapsin I with insulin-containing dense core granules. Thus, although this study establishes that synapsin I is a substrate for CaM kinase II in the pancreatic beta-cell, this event appears not to be important for the mobilization of insulin granules.
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PMID:Site-specific phosphorylation of synapsin I by Ca2+/calmodulin-dependent protein kinase II in pancreatic betaTC3 cells: synapsin I is not associated with insulin secretory granules. 1007 49

The transcription factor CREB is involved in mediating many of the long-term effects of activity-dependent plasticity at glutamatergic synapses. Here, we show that activation of NMDA receptors and voltage-sensitive calcium channels leads to CREB-mediated transcription in cortical neurons via a mechanism regulated by CREB-binding protein (CBP). Recruitment of CBP to the promoter is not sufficient for transactivation, but calcium influx can induce CBP-mediated transcription via two distinct transactivation domains. CBP-mediated transcription is stimulus strength-dependent and can be induced by activation of CaM kinase II, CaM kinase IV, and protein kinase A, but not by activation of the Ras-MAP kinase pathway. These observations indicate that CBP can function as a calcium-sensitive transcriptional coactivator that may act as a regulatory switch for glutamate-induced CREB-mediated transcription.
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PMID:Regulation of CBP-mediated transcription by neuronal calcium signaling. 1023 Jul 99

After cessation of repeated, intermittent amphetamine, we detected an emergent Ca2+-dependent component of amphetamine-induced dopamine release and an increase in calmodulin and Ca2+- and calmodulin-dependent protein kinase activity in rat striatum. This study examined the involvement of calmodulin-dependent protein kinase II (CaM kinase II) and synaptic vesicles in the enhanced Ca2+-dependent dopamine release in response to amphetamine or K+ in rat striatum. Rats were pretreated for 5 d with 2.5 mg/kg amphetamine or saline and withdrawn from drug for 10 d. The selective CaM kinase II inhibitor KN-93 (1 microM), but not the inactive analog KN-92, attenuated the Ca2+-dependent amphetamine-mediated dopamine release from amphetamine-pretreated rats but had no effect in saline-pretreated controls. [3H]Dopamine uptake was unaltered by repeated amphetamine or KN-93 and was Ca2+ independent. Striatal dopamine release stimulated by 50 mM KCl was enhanced twofold after repeated amphetamine compared with that in saline controls but was unaffected by KN-93. To examine the requirement for dopaminergic vesicles in the Ca2+-dependent dopamine release, we administered reserpine to saline- and amphetamine-pretreated rats 1 d before killing. Reserpine pretreatment did not affect amphetamine-mediated dopamine release from either pretreatment group but completely ablated K+-mediated dopamine release. Reserpine did not disrupt the ability of 1 microM KN-93 to block the Ca2+-dependent amphetamine-mediated dopamine release from amphetamine-pretreated rats. The results indicate that the enhanced dopamine release elicited by amphetamine from chronically treated rats is dependent on Ca2+- and calmodulin-dependent phosphorylation and is independent of vesicular dopamine storage. On the contrary, the enhanced depolarization-mediated vesicular dopamine release is independent of Ca2+- and calmodulin-dependent phosphorylation.
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PMID:Enhanced amphetamine- and K+-mediated dopamine release in rat striatum after repeated amphetamine: differential requirements for Ca2+- and calmodulin-dependent phosphorylation and synaptic vesicles. 1023 12


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