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

Endogenous phosphorylation of the crude membrane fraction of cultured 3Y1 fibroblast cells was enhanced by the addition of Ca2+/calmodulin. Both Ca2+/calmodulin-dependent protein kinase activity and its substrate were present in a cytoskeletal fraction, obtained as a pellet after washing of the membrane fraction with 2 mM EGTA, 0.6 M NaCl, and 1% Triton X-100. The phosphorylatable protein in the Triton X-insoluble fraction was identified by immunoblotting as vimentin. This endogenous phosphorylation induced by calmodulin was inhibited by the addition of KN-62, a specific Ca2+/calmodulin-dependent protein kinase II inhibitor, in a dose-dependent manner. However, phosphorylation of the 59 kDa protein (vimentin) in this fraction was not stimulated by adding both phosphatidyl serine and cAMP, thereby suggesting the absence of protein kinase C or of cAMP-dependent protein kinase in this fraction. The protein kinase associated with the Triton X-insoluble fraction phosphorylated the Ca2+/calmodulin-dependent protein kinase II-specific site of synapsin I from the bovine cortex. Two-dimensional phosphopeptide maps of vimentin indicated that a major phosphopeptide phosphorylated by the endogenous calmodulin-dependent kinase also appears to be the same as a major phosphopeptide phosphorylated by the exogenous Ca2+/calmodulin-dependent protein kinase II. Our results suggest that cytoskeleton-associated Ca2+/calmodulin-dependent protein kinase II regulates dynamic cellular functions through the phosphorylation of cytoskeletal elements in non-neural cells.
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PMID:Ca2+/calmodulin-dependent protein phosphorylation associated with the cytoskeleton of quiescent rat fibroblast (3Y1) cells. 166 12

In previous studies, we described a soluble Ca2+/calmodulin-dependent protein kinase which is the major Ca2+/calmodulin-dependent microtubule-associated protein 2 (MAP-2) kinase in rat brain [Schulman, H. (1984) J. Cell Biol. 99, 11-19; Kuret, J. A., & Schulman, H. (1984) Biochemistry 23, 5495-5504]. We now demonstrate that this protein kinase has broad substrate specificity. Consistent with a multifunctional role in cellular physiology, we show that in vitro the enzyme can phosphorylate numerous substrates of both neuronal and nonneuronal origin including vimentin, ribosomal protein S6, synapsin I, glycogen synthase, and myosin light chains. We have used MAP-2 to purify the enzyme from rat lung and show that the brain and lung kinases have nearly indistinguishable physical and biochemical properties. A Ca2+/calmodulin-dependent protein kinase was also detected in rat heart, rat spleen, and in the ring ganglia of the marine mollusk Aplysia californica. Partially purified MAP-2 kinase from each of these three sources displayed endogenous phosphorylation of a 54 000-dalton protein. Phosphopeptide analysis reveals a striking homology between this phosphoprotein and the 53 000-dalton autophosphorylated subunit of the major rat brain Ca2+/calmodulin-dependent protein kinase. The enzymes phosphorylated MAP-2, synapsin I, and vimentin at peptides that are identical with those phosphorylated by the rat brain kinase. This enzyme may be a multifunctional Ca2+/calmodulin-dependent protein kinase with a widespread distribution in nature which mediates some of the effects of Ca2+ on microtubules, intermediate filaments, and other cellular constituents in brain and other tissues.
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PMID:Ca2+/calmodulin-dependent microtubule-associated protein 2 kinase: broad substrate specificity and multifunctional potential in diverse tissues. 407 98

Hydrolysis of inositol phospholipids by receptor stimulation activates two separate signaling pathways, one leading to the activation of protein kinase C (C kinase) via formation of diacylglycerol. The other is the inositol trisphosphate (IP3)/Ca2+ pathway and a major downstream kinase which is activated is Ca2+/calmodulin-dependent protein kinase II (CaM kinase II). To examine signaling pathways of C kinase and CaM kinase II to the cytoskeletal protein vimentin, we prepared monoclonal antibodies YT33 and MO82 which recognize the phosphorylation state of vimentin by C kinase and by CaM kinase II, respectively. Ectopic expression of constitutively active C kinase or CaM kinase II in primary cultured astrocytes by microinjection of the corresponding expression vectors induced phosphorylation of vimentin at each specific phosphorylation site, followed by reorganization of vimentin filament networks. In contrast, simultaneous activation of C kinase and CaM kinase II by inositol phospholipid hydrolysis with receptor stimulation led to an exclusive phosphorylation of vimentin at the CaM kinase II site, not at the site of C kinase. These results indicate that the intracellular targeting of C kinase and CaM kinase II signalings to vimentin is regulated separately, under physiological conditions.
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PMID:Differential targeting of protein kinase C and CaM kinase II signalings to vimentin. 749 Feb 82

We investigated the activation of Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) via stimulation of glutamate receptors and subsequent phosphorylation of vimentin and glial fibrillary acidic protein (GFAP) in cultured rat cortical astrocytes. The indirect immunofluorescence analysis with the anti-CaM kinase II antibody revealed that the enzyme was detected diffusely in the cytoplasm and more intensely in the nucleus. Glutamate elevated the Ca(2+)-independent activity of CaM kinase II through autophosphorylation, and this response was blocked by both DL-2-amino-3-phosphonopropionate and 6-cyano-7-nitroquinoxaline-2,3-dione, but not by D-2-amino-5-phosphonovalerate. In the experiments using 32P-labeled astrocytes, the phosphorylation of vimentin and GFAP as well as autophosphorylation of CaM kinase II were found to be stimulated after the exposure to glutamate. It was concluded by two-dimensional phosphopeptide analysis that the increased phosphorylation of vimentin and GFAP observed in intact cells were due to the activation of CaM kinase II by glutamate. These results suggest that glutamate can activate CaM kinase II through stimulation of both the metabotropic and non-N-methyl-D-aspartate receptors, and that the concomitant phosphorylation of vimentin and GFAP may in turn regulate the functions of intermediate filament proteins in intact astrocytes.
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PMID:Activation of Ca2+/calmodulin-dependent protein kinase II and phosphorylation of intermediate filament proteins by stimulation of glutamate receptors in cultured rat cortical astrocytes. 790 75

Ca2+ plays a central role in cell signaling, and Ca2+/calmodulin-dependent protein kinase II (CaMKII) is a major mediator of Ca2+ actions. The spatial distribution of intracellular Ca2+ signaling is not homogenous, rather it is dynamically organized, and it has been speculated that spatial patterns of Ca2+ signals may function as a form of cellular information transmitted to downstream molecules. To address this issue, we studied the intracellular distributions of the signalings by CaMKII and Ca2+ in the same astrocytes. The former was visualized by monitoring site-specific phosphorylation of a cytoskeletal protein vimentin, using site- and phosphorylation-specific antibodies, while the latter was examined by fura-2-based Ca2+ microscopy. Local Ca2+ signals induced vimentin phosphorylation by CaMKII localized in the same area. On the other hand, Ca2+ waves in astrocytes induced global phosphorylation of vimentin by CaMKII. A small population of vimentin filaments highly phosphorylated by CaMKII underwent structural alteration into short filaments at electron microscopic level. These results indicate that CaMKII transmits spatial patterns of Ca2+ signals to vimentin as cellular information. The possibility is discussed that spatial patterns of vimentin phosphorylation may be important for intracellular organization of vimentin filament networks.
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PMID:Spatial patterns of Ca2+ signals define intracellular distribution of a signaling by Ca2+/Calmodulin-dependent protein kinase II. 931 33

It has been shown that cells with high affinity very late Ag (VLA)-integrins have up-regulated expression of a beta1-subunit epitope, which is detected by 15/7 mAb. In this study, we demonstrate that soluble VCAM-1 (sVCAM-1) exhibits chemotactic activity of T cells with high affinity VLA-4 against VCAM-1, such as Jurkat T cells and IL-2-dependent T cells. Moreover, we found that T cells in the synovial fluid show high basal migration in the absence of sVCAM-1, compared with peripheral blood T cells in patients with rheumatoid arthritis. Among T cells in the synovial fluid, CD45RO+ memory T cells, in response to sVCAM-1, showed a much higher than basal migratory response when compared with CD45RA+ naive cells, while no significant difference was observed between CD4+ and CD8+ T cells. The chemotactic activity of sVCAM-1 is inhibited in the presence of anti-VCAM-1 and anti-VLA-4, which interfered with the binding between VCAM-1 and VLA-4. Inhibition studies using various kinase inhibitors (C3 exoenzyme, KN62, and H7) show that Rho, Ca2+/calmodulin-dependent kinase II, and protein kinase C are involved in signal transduction in sVCAM-1-induced chemotaxis, respectively, whereas tyrosine kinase seems to play a lesser role, since genistein showed only partial inhibition of T cell chemotaxis. Western blot analysis using an anti-phospho-serine mAb (MO82) reveals that Ser82 in the vimentin is phosphorylated specifically by Ca2+/calmodulin-dependent kinase II through sVCAM-1 activation in the IL-2 dependent T cells. Collectively, by inducing migration and recruitment of T cells through several kinase activations, sVCAM-1 contributes to the development of the inflammation of synovial lesion.
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PMID:Soluble VCAM-1 induces chemotaxis of Jurkat and synovial fluid T cells bearing high affinity very late antigen-4. 979 28

A single dose of diisopropyl phosphorofluoridate (DFP), an organophosphorus ester, produces delayed neurotoxicity (OPIDN) in hen. DFP produces mild ataxia in hens in 7-14 days, which develops into severe ataxia or paralysis as the disease progresses. Since, OPIDN is associated with alteration in the expression of several proteins (e.g., Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) alpha-subunit, tau, tubulin, neurofilament (NF) protein, vimentin, GFAP) as well as their mRNAs (e.g., NF, CaM kinase II alpha-subunit), we determined the effect of a single dose of DFP on the expression of one of the best known immediate-early gene (IEG), c-fos. C-fos expression was measured by Northern hybridization in cerebrum, cerebellum, brainstem, midbrain, spinal cord, and the sciatic nerves of hens at 0.5 hr, 1 hr, 2 hr, 1 day, 5 days, 10 days, and 20 days after a single 1.7 mg/kg, sc. injection of DFP. All the tissues (cerebrum, 52%; cerebellum, 55%; brainstem, 49%; midbrain, 23%; spinal cord, 80%; sciatic nerve, 157%) showed significant increase in c-fos expression in 30 min and this elevated level persisted at least up to 2 hr. Expressions of beta-actin mRNA and 18S RNA were used as internal controls. The significant increase in c-fos expression in DFP-treated hens suggests that c-fos may be one of the IEGs involved in the development of OPIDN.
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PMID:C-fos mRNA induction in the central and peripheral nervous systems of diisopropyl phosphorofluoridate (DFP)-treated hens. 1076 75

Diisopropyl phosphorofluoridate (DFP) is a type I organophosphorus compound and produces delayed neurotoxicity (OPIDN) in adult hens. A single dose of DFP (1.7 mg/kg, s.c.) produces mild ataxia in hens in 7-14 days, which develops into severe ataxia or paralysis as the disease progresses. We have previously shown altered expression of several proteins (e.g. Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) alpha-subunit, tau, tubulin, neurofilament protein (NF), vimentin, GFAP) and an immediate early gene (e.g. c-fos) in DFP-treated hens. Here we show an increase in protein kinase A (PKA) protein level and activity in the spinal cord at 1-day and 5-days time periods after DFP administration. We also determined the protein levels of protein kinase C (PKC), CaM kinase II and several phosphatases (i.e. phosphatase 1 (PP1), phosphatase 2A (PP2A), phosphatase 2B (PP2B) in the spinal cord of DFP-treated hens after 1, 5, 10, and 20 days). There was increase in CaM kinase II alpha subunit level after 10 and 20 days of treatment, and decrease in PKC level at 1-day and 20-days time periods in spinal cord mitochondria. In contrast, the cerebrum, which is resistant to DFP-induced axonal degeneration, did not show change in PKA and CaM Kinase II levels at any time period DFP post-administration. No alteration was found in the protein levels of PP1, PP2A, and PP2B at any time period. An early induction in PKA, which is an important protein kinase in signal transduction, followed by that of CaM kinase might be contributing towards the development of OPIDN in DFP-treated hens.
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PMID:Enhanced activity and level of protein kinase A in the spinal cord supernatant of diisopropyl phosphorofluoridate (DFP)-treated hens. Distribution of protein kinases and phosphatases in spinal cord subcellular fractions. 1145 76

The regulation of the multifunctional calcium/calmodulin dependent protein kinase II (CaMKII) by serine/threonine protein phosphatases has been extensively studied in neuronal cells; however, this regulation has not been investigated previously in fibroblasts. We cloned a cDNA from SV40-transformed human fibroblasts that shares 80% homology to a rat calcium/calmodulin-dependent protein kinase phosphatase that encodes a PPM1F protein. By using extracts from transfected cells, PPM1F, but not a mutant (R326A) in the conserved catalytic domain, was found to dephosphorylate in vitro a peptide corresponding to the auto-inhibitory region of CaMKII. Further analyses demonstrated that PPM1F specifically dephosphorylates the phospho-Thr-286 in autophosphorylated CaMKII substrate and thus deactivates the CaMKII in vitro. Coimmunoprecipitation of CaMKII with PPM1F indicates that the two proteins can interact intracellularly. Binding of PPM1F to CaMKII involves multiple regions and is not dependent on intact phosphatase activity. Furthermore, overexpression of PPM1F in fibroblasts caused a reduction in the CaMKII-specific phosphorylation of the known substrate vimentin(Ser-82) following induction of the endogenous CaM kinase. These results identify PPM1F as a CaM kinase phosphatase within fibroblasts, although it may have additional functions intracellularly since it has been presented elsewhere as POPX2 and hFEM-2. We conclude that PPM1F, possibly together with the other previously described protein phosphatases PP1 and PP2A, can regulate the activity of CaMKII. Moreover, because PPM1F dephosphorylates the critical autophosphorylation site of CaMKII, we propose that this phosphatase plays a key role in the regulation of the kinase intracellularly.
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PMID:Regulation of the multifunctional Ca2+/calmodulin-dependent protein kinase II by the PP2C phosphatase PPM1F in fibroblasts. 1514 Aug 79

Cytosolic phospholipase A2alpha (cPLA2alpha) preferentially hydrolyzes phospholipids containing arachidonic acid and plays a key role in the biosynthesis of eicosanoids. This review discusses the essential features of cPLA2alpha regulation and addresses new insights into the functional properties of this enzyme. Full activation of the enzyme requires Ca2+ binding to an N-terminal C2 domain and phosphorylation on serine residues. Ca2+ binding induces translocation of cPLA2alpha from the cytosol to the perinuclear membranes. Serine phosphorylation is mediated by mitogen-activated protein kinases (MAPKs), Ca2+/calmodulin-dependent protein kinase II, and MAPK-interacting kinase Mnk1. Interaction with proteins and lipids, which include vimentin, annexins, NADPH oxidase, phosphatidylcholine, phosphatidylinositol 4,5-bisphosphate (PIP2), and ceramide-1-phosphate, can also modulate the activity of cPLA2alpha. Recent evidence has established the physiological and pathological roles of cPLA2alpha using cPLA2alpha knockout mice. This enzyme has been implicated in fertility, striated muscle growth, renal concentration, postischemic brain injury, arthritis, inflammatory bone resorption, intestinal polyposis, pulmonary fibrosis, acute respiratory distress syndrome, and autoimmune encephalomyelitis. Now novel three paralogs, cPLA2beta, cPLA2gamma, and cPLA2delta, have been identified in humans. cPLA2gamma is distinct from others in that it is farnesylated and lacks the C2 domain. Biological roles for these new enzymes have not yet been defined.
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PMID:Regulatory mechanism and physiological role of cytosolic phospholipase A2. 1530 15


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