anisomycin and Glioma

High-grade gliomas (HGG) are the most common brain tumors, with an average survival time of 14 months. A glioma-CpG island methylator phenotype (G-CIMP), associated with better clinical outcome, has been described in low and high-grade gliomas. Mutation of IDH1 is known to drive the G-CIMP status. In some cases, however, the hypermethylation phenotype is independent of IDH1 mutation, suggesting the involvement of other mechanisms. Here, we demonstrate that DNMT1 expression is higher in low-grade gliomas compared to glioblastomas and correlates with phosphorylated c-Jun. We show that phospho-c-Jun binds to the DNMT1 promoter and causes DNA hypermethylation. Phospho-c-Jun activation by Anisomycin treatment in primary glioblastoma-derived cells attenuates the aggressive features of mesenchymal glioblastomas and leads to promoter methylation and downregulation of key mesenchymal genes (CD44, MMP9 and CHI3L1). Our findings suggest that phospho-c-Jun activates an important regulatory mechanism to control DNMT1 expression and regulate global DNA methylation in Glioblastoma.

Topics: Anisomycin; Brain Neoplasms; Cell Line, Tumor; DNA (Cytosine-5-)-Methyltransferase 1; DNA Methylation; Epigenesis, Genetic; Female; Gene Expression Regulation, Neoplastic; Genome, Human; Glioma; Humans; Phosphorylation; Prognosis; Promoter Regions, Genetic; Proto-Oncogene Proteins c-jun; Survival Analysis; Up-Regulation

The promoter of the early growth response gene (Egr-1) has been described to be activated by ionizing radiation, and it seems to be clear that this process involves different mitogen activated protein (MAP) kinases, dependent on the specific cell type examined. However, early steps leading to activation of the corresponding pathways and thus to overexpression of Egr-1 are not well understood. In this study, deletion mutants of the 5' upstream region of the Egr-1 gene were generated which allowed us to correlate the radiation-induction of the Egr-1 promoter in U87 glioma cells to five serum response elements. Based on the data shown, a possible role of two cAMP responsive elements for radiation-dependent promoter regulation could be ruled out. On the basis of activator/inhibitor studies applying fetal bovine serum, EGF, PD98059, anisomycin, SB203580, forskolin and wortmannin, it could be demonstrated that in U87 cells the ERK1/2 and potentially SAPK/JNK, but not the p38MAPK/SAPK2, pathway contribute to the radiation-induction of Egr-1 promoter. In addition, it was observed that irradiated cells secrete a diffusible factor into the culture media which accounts for the radiation-induced promoter upregulation. By blocking growth factor receptor activation with suramin, this effect could be completely abolished.

Topics: Androstadienes; Anisomycin; Blotting, Western; Colforsin; DNA-Binding Proteins; Early Growth Response Protein 1; Epidermal Growth Factor; Flavonoids; Glioma; Humans; Imidazoles; Immediate-Early Proteins; Mitogen-Activated Protein Kinases; p38 Mitogen-Activated Protein Kinases; Promoter Regions, Genetic; Pyridines; Suramin; Transcription Factors; Wortmannin

We previously showed that the aggregated form of Hsp27 in cultured cells becomes dissociated as a result of phosphorylation with various types of stress. In order to clarify the signal transduction cascade involved, the effects of various inhibitors of protein kinases and dithiothreitol on the dissociation of Hsp27 were here examined by means of an immunoassay after fractionation of cell extracts by sucrose density gradient centrifugation. The dissociation of Hsp27 induced by exposure of U251 MG human glioma cells to metals (NaAsO2 and CdCl2), hypertonic stress (sorbitol and NaCI), or anisomycin, an activator of p38 mitogen-activated protein (MAP) kinase, was completely suppressed by the presence of SB 203580 or PD 169316, inhibitors of p38 MAP kinase, but not by PD 98059 and Uo 126, inhibitors of MAP kinase kinase (MEK), nor by staurosporine, Go 6983, and bisindolylmaleimide I, inhibitors of protein kinase C. Phorbol ester (PMA)-induced dissociation of Hsp27 was completely suppressed by staurosporine, Go 6983, or bisindolylmaleimide I and partially suppressed by SB 203580, or PD 169316 but not by PD 98059 or Uo 126, indicating mediation by 2 cascades. The presence of 1 mM dithiothreitol in the culture medium during exposure to chemicals suppressed the dissociation of Hsp27 induced by arsenite and CdCl2 but not by other chemicals. These results suggest that the phosphorylation of Hsp27 is catalyzed by 2 protein kinases, p38 MAP kinase-activated protein (MAPKAP) kinase-2/3 and protein kinase C. In addition, metal-induced signals are sensitive to reducing power.

Topics: Anisomycin; Arsenites; Butadienes; Cadmium Chloride; Carcinogens; Dithiothreitol; Enzyme Inhibitors; Flavonoids; Glioma; Heat-Shock Proteins; Humans; Hydrogen Peroxide; Imidazoles; MAP Kinase Kinase Kinases; MAP Kinase Signaling System; Mitogen-Activated Protein Kinases; Nitriles; Osmotic Pressure; Oxidants; p38 Mitogen-Activated Protein Kinases; Protein Kinase C; Protein Synthesis Inhibitors; Pyridines; Sodium Compounds; Sorbitol; Staurosporine; Tetradecanoylphorbol Acetate; Tumor Cells, Cultured