TOP2B Enzymatic Activity on Promoters and Introns Modulates Multiple Oncogenes in Human Gliomas

Abstract

The epigenetic mechanisms involved in transcriptional regulation leading to malignant phenotype in gliomas remains poorly understood. Topoisomerase IIB (TOP2B), an enzyme that decoils and releases torsional forces in DNA, is overexpressed in a subset of gliomas. Therefore, we investigated its role in epigenetic regulation in these tumors.Experimental Design: To investigate the role of TOP2B in epigenetic regulation in gliomas, we performed paired chromatin immunoprecipitation sequencing for TOP2B and RNA-sequencing analysis of glioma cell lines with and without TOP2B inhibition and in human glioma specimens. These experiments were complemented with assay for transposase-accessible chromatin using sequencing, gene silencing, and mouse xenograft experiments to investigate the function of TOP2B and its role in glioma phenotypes.Results: We discovered that TOP2B modulates transcription of multiple oncogenes in human gliomas. TOP2B regulated transcription only at sites where it was enzymatically active, but not at all native binding sites. In particular, TOP2B activity localized in enhancers, promoters, and introns of PDGFRA and MYC, facilitating their expression. TOP2B levels and genomic localization was associated with PDGFRA and MYC expression across glioma specimens, which was not seen in nontumoral human brain tissue. In vivo, TOP2B knockdown of human glioma intracranial implants prolonged survival and downregulated PDGFRA.Conclusions: Our results indicate that TOP2B activity exerts a pleiotropic role in transcriptional regulation of oncogenes in a subset of gliomas promoting a proliferative phenotype.

TOP2B Is Required to Maintain the Adrenergic Neural Phenotype and for ATRA-Induced Differentiation of SH-SY5Y Neuroblastoma Cells

“…( D ) Plot illustrating the relationship between the presence of TOP2B binding peaks and gene expression changes between WT and BKO cells. Genes are categorised according to the evidence for TOP2B binding from inspection of a published glioma ChIP-seq data set [ 62 ] according to the following scheme: none, no peaks observed in the promoter region or gene body; multiple, multiple peaks present; promoter, peak or peaks present in the promoter region; internal with CTCF, TOP2B peak or peaks at a gene-internal position coincident with CTCF binding; and internal, peak or peaks within the gene, not associated with CTCF. ( E ) Profile of epigenetic features, including glioma TOP2B binding activity [ 62 ] for NOTCH1 , profiles for other relevant genes are shown in Fig.…”

“…It is likely that some of these gene expression changes are a direct consequence of loss of TOP2B activity in gene regulatory regions, whereas others may result from indirect effects or a less localised effect on DNA supercoiling. To examine TOP2B binding in the gene regulatory regions of differentially expressed genes such as NOTCH1 , we used an existing TOP2B ChIP-seq data set derived from glioma cell lines TS543 and BT142 [ 62 ] along with publicly available data for CTCF, open chromatin (ATAC-seq), active promoters (H3K4Me3), and promoter/enhancer regions (H3K27Ac), all derived from SH-SY5Y cells or clonally related lines (SK-N-SH or SHEP). Although the TOP2B ChIP-seq data was derived from a different neural cell type (glioma rather than neuroblastoma), inspection of the accompanying expression data revealed that most genes of interest from our RNA-seq analysis were also expressed in the glioma cells.…”

“…This raises the question of whether NOTCH family members are directly regulated by TOP2B. Examination of published glioma ChIP-seq data [ 62 ] revealed TOP2B binding peaks in the regulatory regions or at internal positions of some, but not all genes discussed above. For example, TOP2B binding is present in the promoter regions of both ASCL1 and HES1 as well as NOTCH1 and NOTCH2 , and an internal TOP2B/CTCF binding peak is found in NOTCH3 (Figs.…”

“…S8. Data were plotted using the Integrated Genome Viewer (IGV, Broad Institute) [75] using the following data sources (GEO accession and citation): SK-N-SH CTCF ChIP-seq, GSM2038347; SH-SY5Y ATAC-seq, GSM2700775 [76]; SH-SY5Y H3K27ac, GSM3676080 [77]; GSM2038349; SH-SY5Y H3K4Me3, GSM2419933 [41,42]; SH-EP H3K4Me3, GSM2419932 [41,42]; and TOP2B ChIP-seq data from TS543 and BT142 glioma cell lines in the absence (DMSO) or presence of etoposide (ETO), GSM3904399, GSM3904400, GSM3904401, and GSM3904402 [62] ◂ Fig. 6 Differential expression of ATRA modulated genes in WT and TOP2B null (BKO98) SH-SY5Y cells lines.…”

Importin α as a molecular marker for investigating the microenvironment of micronuclei

“…CCAR2/DBC1 regulates cell survival and death in response to genotoxic and metabolic stresses (Magni et al, 2015). TOP2B is a well-known enzyme that decoils chromatin and releases torsional force into the DNA (Gonzalez-Buendia et al, 2021). Its association with chromatin sites marked by H3K4me3 (Tiwari et al, 2012) and MN (Muciño-Hernández et al, 2023) has also been reported.…”