casein-kinase-ii and Neuroblastoma

Among the post-translational modifications of α-synuclein, phosphorylation has been reported to modulate the protein's nuclear localization, gene-expression and cytotoxicity. However, its effect on the functional performance of dopaminergic-neurons is not known. We aimed to evaluate the effect of siRNA-silencing of casein kinase (CK)2α in SH-SY5Y-cells overexpressing A53T α-synuclein, in alleviating phosphorylated α-synuclein serine129 (pSyn-129)-induced changes in intracellular Ca

Topics: alpha-Synuclein; Casein Kinase II; Dopamine; Dopaminergic Neurons; Down-Regulation; Humans; Neuroblastoma; Reactive Oxygen Species; RNA, Small Interfering

Mitochondrial Ca

Topics: Axon Initial Segment; Casein Kinase II; Homeostasis; Humans; Mitochondria; Neuroblastoma; Neuronal Plasticity

The vast majority of therapeutic approaches tested so far for prion diseases, transmissible neurodegenerative disorders of human and animals, tackled PrP

Topics: Animals; Casein Kinase II; Cell Line, Tumor; Cell Membrane; Cell Survival; Drug Evaluation, Preclinical; Fluorescent Dyes; Gene Expression; Green Fluorescent Proteins; Harmaline; HEK293 Cells; Hematoxylin; Humans; Mice; Neuroblastoma; Prions; PrPC Proteins; Quinacrine; Tacrolimus

γ-Bisabolene has demonstrated antiproliferative activities against several human cancer cell lines. This study first discloses the antiproliferative and apoptosis induction activities of γ-bisabolene to human neuroblastoma TE671 cells. A CC50 value of γ-bisabolene was 8.2 μM to TE671 cells. Cell cycle analysis with PI staining showed γ-bisabolene elevating the sub-G1 fractions in a time-dependent manner. In addition, annexin V-FITC/PI staining showed γ-bisabolene significantly triggering early (annexin-V positive/PI negative) and late (annexin-V positive/PI positive) apoptosis in dose-dependent manners. γ-Bisabolene induced caspase 3/8/9 activation, intracellular ROS increase, and mitochondrial membrane potential decrease in apoptosis of human neuro-blastoma cells. Moreover, γ-bisabolene increased p53 phosphorylation and up-regulated p53-mediated apoptotic genes Bim and PUMA, as well as decreased the mRNA and protein levels of CK2α. Notably, the results indicated the involvement of CK2α-p53 pathways in mitochondria-mediated apoptosis of human neuroblastoma cells treated with γ-bisabolene. This study elucidated the apoptosis induction pathways of γ-bisabolene-treated neuroblastoma cells, in which could be useful for developing anti-neuroblastoma drugs.

Topics: Antineoplastic Agents; Casein Kinase II; Caspases; Cell Line; Cell Proliferation; Cell Survival; Dose-Response Relationship, Drug; Gene Expression Regulation, Neoplastic; Humans; Membrane Potential, Mitochondrial; Neuroblastoma; Sesquiterpenes; Signal Transduction; Tumor Suppressor Protein p53

Ethanol exposure has deleterious effects on the central nervous system. Although several mechanisms for ethanol-induced damage have been suggested, the precise mechanism underlying ethanol-induced neuronal cell death remains unclear. Recent studies indicate that the p75 neurotrophin receptor (p75NTR) has a critical role in the regulation of neuronal survival. This study was designed to examine the role of p75NTR in ethanol-induced apoptotic signaling in neuroblastoma cells. Ethanol caused highly increased level of p75NTR expression. The use of small interfering RNA to inhibit p75NTR expression markedly attenuated ethanol-induced cell cycle arrest and apoptosis. DNA binding activity of Sp1 was increased by ethanol, whereas inhibition of Sp1 activity by mithramycin, a Sp1 inhibitor, or short hairpin RNA suppressed ethanol-induced p75NTR expression. In addition, inhibitors of casein kinase 2 (CK2) and extracellular signal-regulated kinase (ERK) augmented ethanol-induced p75NTR expression. Our results also demonstrate that inhibition of ERK and CK2 caused a further increase in the activation of the p75NTR proximal promoter induced by ethanol. This increased activation was partially suppressed by the deletion of the Sp1 binding sites. These results suggest that Sp1-mediated p75NTR expression is regulated at least in part by ERK and CK2 pathways. The present study also showed that treatment with ethanol resulted in significant increases in the expression of p21, but not the levels of p53 and p53 target genes such as Bax, Puma, and Bcl-2. Furthermore, the inhibition of p75NTR expression or Sp1 activity suppressed ethanol-induced p21 expression, cell cycle arrest, and apoptosis. These data suggest that ethanol increases p75NTR expression, and CK2 and ERK signaling inversely regulate Sp1-mediated p75NTR expression in ethanol-treated neuroblastoma cells. Thus, our study provides more insight into the mechanisms underlying ethanol actions.

Topics: Alcoholism; Apoptosis; Casein Kinase II; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Survival; Central Nervous System Depressants; Cyclin-Dependent Kinase Inhibitor p21; Ethanol; Extracellular Signal-Regulated MAP Kinases; Humans; Nerve Tissue Proteins; Neuroblastoma; Neurons; Receptors, Nerve Growth Factor; Sp1 Transcription Factor; Transcriptional Activation; Tumor Suppressor Protein p53

Acrylamide has been known to have a neurotoxic effect which is associated with nerve damage in both the central and peripheral nervous systems. Since neural cell adhesion molecule (NCAM) plays an important role in the processes of neuronal development and synaptic plasticity, the down-regulation of NCAM may lead impaired spatial memory and reduced long-term potentiation. We examined the effect of acrylamide on NCAM expression and the mechanisms of its effect in human neuroblastoma cells. Treatment with acrylamide resulted in the decrease of NCAM expression, which was reversed by CK2 inhibitor, 4,5,6,7-tetrabromobenzotriazole (TBB). Moreover, Western blot analysis showed that acrylamide induced the expression of CK2. Acrylamide dose-dependently decreased the DNA binding affinity of the Ikaros transcription factor, which is a bifunctional differentiation factor. In addition, the cells treated with acrylamide and CK2 inhibitor showed increased Ikaros activity compared with the acrylamide treatment only. Small interfering RNA-mediated depletion of CK2-α also increased Ikaros activity in acrylamide-treated cells. Overall, these data suggest that acrylamide decreases the Ikaros DNA binding activity via the CK2 pathway, resulting in a decrease of NCAM expression and provide further insight into the mechanisms underlying acrylamide actions.

Topics: Acrylamide; Blotting, Western; Casein Kinase II; Cell Line, Tumor; DNA; Dose-Response Relationship, Drug; Gene Expression Regulation, Neoplastic; Humans; Ikaros Transcription Factor; Neural Cell Adhesion Molecules; Neuroblastoma; Signal Transduction

The antioxidant-activated transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) regulates the induction of cytoprotective genes against chemical toxicity and oxidative injuries. The role of phosphorylation in Nrf2 activation has been suggested but remains elusive. We report that phenolic antioxidant/pro-oxidant tert-butylhydroquinone (tBHQ) induced two forms of the Nrf2 protein in neuroblastoma cells (IMR-32), which migrated as distinctive bands on SDS-PAGE. In vitro treatment with lambda phosphatase eliminated the slower migrating form and increased the amount of the faster migrating form of Nrf2. In vivo (32)Pi-phosphorylation resulted in (32)Pi-labeling of the Nrf2 protein in the presence of tBHQ that can be dephosphorylated by lambda phosphotase, indicating that the slower migrating form is a phosphorylated Nrf2 protein and the faster form an unphosphorylated Nrf2. Unphosphorylated Nrf2 predominated in the cytoplasm, whereas the phosphorylated form preferentially localized in the nucleus. Nuclear Nrf2 can be dephosphorylated by lambda phosphotase in vitro and be converted to the faster migrating form, implicating phosphorylation of Nrf2 in the cytoplasmic-nuclear translocation of the protein. Deletional analyses from both the carboxyl- and amino-ends revealed the transcription activation (TA) domains Neh4 (Nrf2-ECH homology 4) and Neh5 (Nrf2-ECH homology 5) as a major region necessary for the phosphorylation. The TA domains are characterized by the presence of multiple phosphorylation sites of casein kinase 2 (CK2). Moreover, CK2 phosphorylated the TA domains in vitro. Treatment with CK2 inhibitor 2-dimethylamino-4,5,6,7,-tetrabromo-1H-benzimidazole (DMAT) blocked the induction of endogenous target genes of Nrf2 in cells and inhibited the TA activities of both the full length and the TA domains of Nrf2 to a large extent. Finally, phosphorylation of the TA domains correlated with the nuclear translocation of Nrf2 that was inhibited by DMAT in a concentration-dependent manner. The findings demonstrated that phosphorylation of Nrf2 at the TA domains by CK2 is an integral component of Nrf2 activation necessary for the nuclear localization and transcription activation function of Nrf2 in neuroblastoma cells.

Topics: Amino Acid Sequence; Animals; Benzimidazoles; Casein Kinase II; Cell Line, Tumor; Cell Nucleus; Conserved Sequence; Electrophoresis, Polyacrylamide Gel; Humans; Mice; Molecular Sequence Data; Neuroblastoma; NF-E2-Related Factor 2; Phosphorylation; Protein Isoforms; Protein Structure, Tertiary; Protein Transport; Recombinant Proteins; Sequence Deletion; Substrate Specificity; Transcriptional Activation

The distribution of microtubule-associated protein lB (MAPlB) phosphorylated by either proline-directed protein kinase (PDPK) or casein kinase II (CK II) in neuroblastoma cells and hippocampal neurons has been studied by immunofluorescence using specific antibodies to distinct phosphorylation-sensitive epitopes. A proximo-distal gradient of increasing PDPK-catalyzed phosphorylation of MAPlB is superimposed on a proximo distal gradient of decreasing CK II-catalyzed MAPlB phosphorylation within growing axon-like neurites. Additionally, CK II-phosphorylated MAPlB is present in cell bodies and dendrites where no PDPK-phosphorylated MAPlB is observed. These results suggest distinct roles for both types of modifications of MAPlB in developing neurons.

Topics: Animals; Casein Kinase II; Cell Differentiation; Cells, Cultured; Epitopes; Hippocampus; Mice; Microtubule-Associated Proteins; Neurites; Neuroblastoma; Neurons; Phosphorylation; Proline-Directed Protein Kinases; Protein Serine-Threonine Kinases; Rats; Tumor Cells, Cultured

Phosphorylation of microtubule-associated protein MAP1B and the neuronal-specific beta III-tubulin isoform takes place during neurite growth in neuroblastoma cells. Protein kinase CK2 (formerly referred to as casein kinase 2) is possibly involved in beta III-tubulin phosphorylation. As for MAP1B, there are at least two types of phosphorylation; one catalyzed by proline-directed protein kinases and another catalyzed by CK2. Protein kinase CK2 is primarily localized to the nuclei in proliferating neuroblastoma cells, whereas an increased amount of the enzyme is present in the cytoplasm of postmitotic cells bearing neurites. Treatment of neuroblastoma cells with an antisense oligonucleotide which specifically results in CK2 catalytic subunit depletion inhibits neuritogenesis. CK2 depletion is accompanied by dephosphorylation of MAP1B on the corresponding phosphorylatable sites. This dephosphorylation is paralleled by a release of MAP1B from microtubules. These results suggest that MAP1B phosphorylation by CK2 may be required for the assembly of microtubules within neurites. Other neuronal cytoskeletal proteins including MAP1A and tau are also substrates for CK2, indicating a role for the enzyme in the regulation of cytoskeletal functions also in mature neurons.

Topics: Amino Acid Sequence; Animals; Casein Kinase II; Cytoplasm; Epitopes; Mice; Microtubule-Associated Proteins; Molecular Sequence Data; Neoplasm Proteins; Nerve Tissue Proteins; Neurites; Neuroblastoma; Phosphorylation; Protein Processing, Post-Translational; Protein Serine-Threonine Kinases; Tubulin; Tumor Cells, Cultured

1. The use of antisense oligonucleotides to inhibit expression of the genes coding for the catalytic (alpha/alpha') and regulatory (beta) subunits of protein kinase casein kinase 2 (CK2) has allowed study of the role of this enzyme in mouse neuroblastoma cells. 2. Selective depletion of catalytic (alpha/alpha') subunits results in the blocking of neuritogenesis. The depletion of catalytic subunits also affects the sorting of the regulatory (beta) subunit of CK2, as the absence of catalytic subunits prevents the translocation of the regulatory subunit to the nuclei. These results emphasize the existence of a control mechanism linking the expression and sorting of CK2 catalytic and regulatory subunits. 3. Selective depletion of the regulatory (beta) subunit of protein kinase CK2 by an specific antisense oligonucleotide causes partial inhibition of neurite extension.

Topics: Allosteric Site; Animals; Base Sequence; Biological Transport; Casein Kinase II; Catalysis; Cell Differentiation; Cell Nucleus; Enzyme Induction; Gene Expression Regulation, Neoplastic; Mice; Molecular Sequence Data; Neoplasm Proteins; Neurites; Neuroblastoma; Oligonucleotides, Antisense; Protein Conformation; Protein Serine-Threonine Kinases; Protein Structure, Tertiary; Tumor Cells, Cultured

Whereas cells from most clonal lines derived from the murine neuroblastoma C1300 tumor can be induced to differentiate by serum withdrawal from culture medium, the NIA-103 clonal cell line has been considered unable to extend axon-like processes (neurites). Neurite growth depends on microtubule protein assembly, and although NIA-103 cells have essentially the same amounts of microtubule-associated protein (MAP)-1B and the neuronal-specific class beta 3-tubulin isoform as other neuroblastoma cell lines, these proteins are not phosphorylated in NIA-103 cells on serum withdrawal. The lack of microtubule protein phosphorylation may be due to the different sorting between the nucleus and the cytoplasm of the casein kinase II-related enzyme that is possibly involved in the modification of microtubule proteins. It is interesting that addition of DNA synthesis inhibitors to serum-starved NIA-103 cell cultures induces an increase in the level of cytosolic casein kinase II, an augmented in situ phosphorylation of MAP-1B, and the extension of neurites. Thus, the level of cytoplasmic casein kinase II appears to be controlled by the growth status of neuroblastoma cells. The shift to an increased cytoplasmic concentration of casein kinase II in nonproliferating, differentiating neuroblastoma cells is consistent with its putative role in the regulation of the cytoskeletal rearrangements underlying neuronal morphogenesis and plasticity.

Topics: Animals; Casein Kinase II; Cell Differentiation; Cytoplasm; DNA, Neoplasm; Isomerism; Microtubule Proteins; Neurites; Neuroblastoma; Phosphorylation; Protein Serine-Threonine Kinases; Tubulin; Tumor Cells, Cultured