Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: UMLS:C0027819 (neuroblastoma)
27,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have isolated and sequenced cDNAs encoding Ca2+/calmodulin-dependent protein kinase type Gr (CaM-K-Gr, also called CaM-K-IV) from human brain and thymus. The sequence of the protein coding region of the cDNA is identical in both brain and thymus, although Northern hybridization analysis shows variation of the mRNA transcripts in these tissues. The sequence predicts a protein of M(r) 51,897 that is 83.7% identical and shows 89.2% similarity with the rat homologue. The deduced human CaM-K-Gr is identical to the rat and mouse proteins in the portion of the enzyme involved in ATP binding, the catalytic domain and Ca2+/calmodulin-binding domain; however, the N terminus of the human kinase, which may comprise a second regulatory domain [McDonald et al., J. Biol. Chem. 268 (1993) 10054-10059], contains a 4-amino-acid (aa) insertion relative to the rodent enzymes. Additionally, the C-terminal association domain shows only 45.2 and 41.6% identity with the rat and mouse proteins, respectively, suggesting that this domain is not constrained by stringent structural and functional requirements. Based on the predicted aa sequence of the human kinase, we produced polyclonal antisera against a C-terminal peptide that recognizes two forms of CaM-K-Gr in human T-cell lymphoma and neuroblastoma cell lines. The human antiserum cross-reacts with the rat and mouse proteins and immunoprecipitates the active kinase.
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PMID:The cDNA sequence and characterization of the Ca2+/calmodulin-dependent protein kinase-Gr from human brain and thymus. 819 51

Bordetella pertussis produces a calmodulin-stimulated adenylyl cyclase that invades animal cells and raises intracellular cAMP levels. The enzyme does not enter animal cells by receptor-mediated endocytosis, but the mechanism for invasion of animal cells has not been defined. We have proposed that the 177 kDa adenylyl cyclase is proteolyzed to a 45 kDa catalytic subunit and one or more polypeptides (invasive factor) that facilitate entry of the catalytic subunit into animal cells. In this study, we report the identification of a sequence of amino acids within the adenylyl cyclase catalytic subunit that is important for entry of the enzyme into eukaryotic cells. A synthetic peptide corresponding to amino acids 313-339 within the catalytic subunit was shown to inhibit invasion of neuroblastoma cells by the adenylyl cyclase. In addition, this peptide inhibited the association of the catalytic subunit with invasive factor. We propose that this domain is a site for interaction between the catalytic subunit and invasive factor.
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PMID:Identification of a domain in Bordetella pertussis adenylyl cyclase important for subunit interactions and cell invasion activity. 825 8

Effects of the calmodulin inhibitor calmidazolium on stimulation of nitric oxide (NO) release were investigated in neuroblastoma N1E-115 cells. NO release was determined indirectly by measuring cyclic GMP formation. Instead of the expected decrease in NO generation based on the calmodulin dependence of neuronal NO synthase, calmidazoline paradoxically increased cyclic GMP formation. Maximal activation occurred at 3 min and the effects were concentration dependent. This calmidazolium-stimulated NO release was markedly blocked by hemoglobin and N-monomethyl-L-arginine.
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PMID:The calmodulin antagonist calmidazolium stimulates release of nitric oxide in neuroblastoma N1E-115 cells. 838 25

In neuroblastoma x glioma NG108-15 hybrid cells, opioid agonists inhibited both basal and prostaglandin E1-stimulated adenylate cyclase activities assayed in the presence of the phosphodiesterase (PDE) inhibitors isobutylmethylxanthine and ZK62711 (rolipram). However, when intracellular [3H]cAMP was measured in the absence of the PDE inhibitors the maximal inhibitory level was increased, using the opioid agonist D-Ala2,D-Leu5-enkephalin. This increase in opioid activity was due to agonist stimulation of cAMP degradation, because when the degradation rate of [3H] cAMP was measured in intact hybrid cells it was observed to increase from the control value of 0.495 +/- 0.003 min-1 to 0.760 +/- 0.003 min-1 in the presence of 1 microM D-Ala2,D-Leu5-enkephalin; this was reversed by naloxone. Dose-dependent studies with various opioid agonists, partial agonists, and antagonists revealed that there was a direct correlation between the abilities of these opioid ligands to inhibit adenylate cyclase activity and to stimulate PDE activity, with enkephalin and its analogs being the most potent agonists. Chronic agonist treatment also resulted in a reduction of the opioid agonist stimulation of cAMP degradation, with an apparent decrease in the PDE activity upon addition of naloxone after chronic treatment. However, treatment of the hybrid cells with pertussis toxin, which attenuated the agonist inhibition of adenylate cyclase activity, did not abolish this opioid response. When selective inhibitors for various types of PDE were used, the type I PDE inhibitor W-7 attenuated the opioid effect, whereas the type II PDE inhibitor trequinsin (HL725), the type III PDE inhibitor indolidan, and the type IV PDE inhibitor rolipram had no effect on opioid-stimulated cAMP degradation. The stimulation of type I PDE activity by delta-opioid receptors was independent of extracellular Ca2+ and was not observed with membrane preparations. Therefore, in NG108-15 cells delta-opioid receptors regulate intracellular cAMP levels by coupling to a pertussis toxin-insensitive guanine nucleotide-binding protein, resulting in an increase in intracellular Ca2+ and in Ca2+/calmodulin-dependent PDE activity.
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PMID:delta-Opioid receptor activates cAMP phosphodiesterase activities in neuroblastoma x glioma NG108-15 hybrid cells. 838 86

Characterization of 'low Km' 3':5' cyclic nucleotide phosphodiesterase activities (PDE) expressed in mouse N18TG2 neuroblastoma cells is reported. At least 3 peaks of activity were isolated by DEAE chromatography, none of which was calcium-calmodulin stimulated and cGMP stimulated or inhibited. A first peak elutes at 200 mM sodium acetate; it specifically hydrolyzes cGMP with a Km of 4.7 microM and shows sensitivity to zaprinast [M&B 22948] (1.8 microM). A second peak eluting at 410 mM sodium acetate hydrolyzes both cyclic nucleotides. A third peak, specific for cAMP hydrolysis, elutes at 580 mM sodium acetate, has a Km of 3.2 microM and is sensitive to RO 20 1724 (7.6 microM) and rolipram (2 microM). Hydrodynamic analysis showed for the first peak a Stokes radius of 5.3 nm with a sedimentation coefficient of 8.1 S, a frictional ratio (f/fo) of 1.41 and a native molecular mass of 182 kDa. The same analysis for peak 3 showed a Stokes radius of 4.1 nm with a sedimentation coefficient of 3.2 S, a frictional ratio of 1.63 and a native molecular mass of 56 kDa. The biochemical features reported for the enzyme eluting in the first peak, and its cGMP-binding activity stimulated by inhibitors of phosphodiesterase activity, demonstrate that it belongs to the PDE V subfamily; on the other hand the cAMP specific enzyme eluting in the third peak can be assigned to the 'RO 20 1724 inhibited' form. The significance of these findings is discussed in relation to the functional characteristics of the N18TG2 cell line.
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PMID:Characterization of 3':5' cyclic nucleotide phosphodiesterase activities of mouse neuroblastoma N18TG2 cells. 838 2

Diethyldithiocarbamate (DDC) was used to treat the neuroblastoma cell line Neuro-2a. Cell injury caused by DDC affects the calcium-binding protein calmodulin (CaM) and alters copper homeostasis in these cells. Neuro-2a cells were treated with 1 x 10(-5) M DDC for 1 h and were harvested at various time points over a 24-h period. Light microscopy of control cells showed CaM deposited around the cell periphery and along the neuritic processes. Treated cells showed the same distribution until 3 h after treatment. Electron microscopy showed CaM deposited around the cell periphery and within the cytoplasm and nucleus of control cells. Treated cells showed a time-dependent localization of CaM in relation to cellular disorganization. Staining of electrophoretic transfers by ProtoGold showed that CaM was present in all control samples and treated samples through 6 h. Atomic absorption spectrophotometry showed no difference in calcium levels between control and treated samples, but copper levels were significantly elevated. This study indicated that degenerative changes induced by DDC altered calmodulin levels. These changes may have been caused by elevated copper content within the cells and subsequent cell injury.
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PMID:Effects of diethyldithiocarbamate on calmodulin in neuroblastoma cells. 839 42

An 80-kDa protein labeled with [3H]myristic acid in C6 glioma and N1E-115 neuroblastoma cells has been identified as the myristoylated alanine-rich C kinase substrate (MARCKS protein) on the basis of its calmodulin-binding, acidic nature, heat stability, and immunochemical properties. When C6 cells preincubated with [3H]myristate were treated with 200 nM 4 beta-12-O-tetradecanoylphorbol 13-acetate (beta-TPA), labeled MARCKS was rapidly increased in the soluble digitonin fraction (maximal, fivefold at 10 min) with a concomitant decrease in the Triton X-100-soluble membrane fraction. However, phosphorylation of this protein was increased in the presence of beta-TPA to a similar extent in both fractions (maximal, fourfold at 30 min). In contrast, beta-TPA-stimulated phosphorylation of MARCKS in N1E-115 cells was confined to the membrane fraction only and no change in the distribution of the myristoylated protein was noted relative to alpha-TPA controls. These results indicate that although phosphorylation of MARCKS by protein kinase C occurs in both cell lines, it is not directly associated with translocation from membrane to cytosol, which occurs in C6 cells only. The cell-specific translocation of MARCKS appears to correlate with previously demonstrated differential effects of phorbol esters on stimulation of phosphatidylcholine turnover in these two cell lines.
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PMID:Dissociation of phosphorylation and translocation of a myristoylated protein kinase C substrate (MARCKS protein) in C6 glioma and N1E-115 neuroblastoma cells. 845 32

The roles of protein kinases and calmodulin in regulating neurite outgrowth in murine neuroblastoma NS-20Y cells were investigated by testing the effect of various inhibitors on the neuritogenesis induced by serum deprivation. The percentage of cells with neurites was low (1-3%) in medium containing 10% serum, but reached about 50-60% when the cells were cultured for 24 h in serum-free medium. W-7 (10 microM), calmidazolium (0.3 microM), and trifluoperazine (0.1 microM), drugs reported to inhibit calmodulin-dependent events, reduced neurite outgrowth. On the other hand, H-7 (inhibitor of protein kinase C and cyclic AMP-dependent protein kinase) and H-89 (inhibitor of cyclic AMP-dependent protein kinase) were ineffective. Genistein (inhibitor of protein tyrosine kinase) and wortmannin (inhibitor of phosphatidylinositol 3-kinase) did not affect the number of cells with neurites. Activation of protein kinases, which is blocked by these inhibitors, does not appear to be essential to the extension and maintenance of neurites. KN-62 and KN-93 (inhibitors of Ca2+/calmodulin-dependent protein kinase II) were also tested but did not inhibit neurite outgrowth. These results suggest that a calmodulin-dependent process, other than the activation of Ca2+/calmodulin-dependent protein kinase II, is involved in the neuritogenesis in murine neuroblastoma NS-20Y cells in serum-free medium.
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PMID:Inhibition of neurite outgrowth in murine neuroblastoma NS-20Y cells by calmodulin inhibitors. 854 72

Because changes in intracellular Ca2+ affect progression through the mitotic cell cycle, we investigated the role of Ca2+-binding proteins in regulating cell cycle progression. Evidence was found demonstrating that the activation of Ca2+/calmodulin-dependent protein kinase (CaM kinase) inhibits cell cycle progression in small cell lung carcinoma (SCLC) cells. We also demonstrated that SCLC cells express both CaM kinase type II (CaMKII) and CaM kinase type IV (CaMKIV). Five independent SCLC cell lines expressed proteins reactive with antibody to the CaMKII beta subunit, but none expressed detectable proteins reactive with antibody to the CaMKII alpha subunit. All SCLC cell lines tested expressed both the alpha and beta isoforms of CaMKIV. Immunoprecipitation of CaMKII from SCLC cells yielded multiple proteins that autophosphorylated in the presence of Ca2+ / calmodulin. Autophosphorylation was inhibited by the CaMKII(281-302) peptide, which corresponds to the CaMKII autoinhibitory domain, and by 1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4- phenylpiperazine (KN-62), a specific CaM kinase antagonist. Influx of Ca2+ through voltage-gated Ca2+ channels stimulated phosphorylation of CaMKII in SCLC cells, and this was inhibited by KN-62. Incubation of SCLC cells of KN-62 potently inhibited DNA synthesis, and slowed progression through S phase. Similar anti-proliferative effects of KN-62 occurred in SK-N-SH human neuroblastoma cells, which express both CaMKII and CaMKIV, and in K562 human chronic myelogenous leukemia cells, which express CaMKII but not CaMKIV. The expression of both CaMKII and CaMKIV by SCLC cells, and the sensitivity of these cells to the anti-proliferative effects of KN-62, suggest a role for CaM kinase in regulating SCLC proliferation.
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PMID:Expression of Ca2+/calmodulin-dependent protein kinase types II and IV, and reduced DNA synthesis due to the Ca2+/calmodulin-dependent protein kinase inhibitor KN-62 (1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenyl piperazine) in small cell lung carcinoma. 861 9

Following mobilization with the inositol 1,4,5-trisphosphate (IP3)-generating agonist bradykinin, Ca2+ stores in neuroblastoma x glioma hybrid, NG108-15 cells require extracellular Ca2+ to refill. The process by which this store refills with Ca2+ was characterized by recording bradykinin-induced intracellular free Ca2+ concentration transients as an index of the degree of refilling of the store. Cyclopiazonic acid, a microsomal Ca2+ ATPase inhibitor, reversibly depleted intracellular Ca2+ stores in these cells, but did not recruit detectable Ca2+ influx, suggesting that these cells lack substantial capacitative Ca2+ entry. The paucity of voltage-sensitive Ca2+ channels in undifferentiated NG108-15 cells, suggested that a channel analogous to that proposed to mediate capacitative Ca2+ entry in nonexcitable cells might assist refilling IP3-sensitive Ca2+ stores in these cells. The possibility that compounds shown previously to inhibit capacitative Ca2+ entry in nonexcitable cells might inhibit the refilling of the IP3-sensitive store in NG108-15 cells was explored. The IP3-sensitive store was depleted by exposure to bradykinin, allowed to refill briefly in the presence of the test compound and then challenged again with bradykinin to evaluate the degree of refilling of the store. The imidazole derivatives, econazole (10 microM), L-651582 (10 microM) and SKF 96365 (20 microM), all completely blocked the bradykinin-induced Ca2+ response. Calmodulin antagonists, W-7 (100 microM) and trifluoperazine (10 microM), were also effective, although at concentrations well above those required to inhibit calmodulin. Because of the high concentrations required to inhibit bradykinin responses, the possibility that these agents might have additional effects was explored. Compounds were tested in a paradigm in which the store was preloaded with Ca2+ before treatment. All of these agents depleted, at least partially, the preloaded store. Econazole was the least effective of the compounds tested for releasing stores, although it was comparable to the other compounds for inhibition of refilling. Although NG108-15 cells refill intracellular Ca2+ stores by a plasmalemmal Ca2+ leak, this leak shares a pharmacology similar to the capacitative Ca2+ entry pathway described for nonexcitable cells.
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PMID:Pharmacologic characterization of refilling inositol 1,4,5-trisphosphate-sensitive Ca2+ stores in NG108-15 cells. 875 Sep 56


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