Mapping catalytically engaged TOP2B in neurons reveals the principles of topoisomerase action within the genome [Re-analysis_TOP2B_mm10]

Topoisomerase IIb (TOP2B) is essential for neural development and function, yet its precise molecular roles remain poorly understood. Here we trapped catalytically engaged TOP2B in DNA cleavage complexes (TOP2Bccs) and mapped their positions genome-wide in cultured mouse cortical neurons. We report that chromosome compartments set the threshold of TOP2B activity within the genome, while specific nucleosome configurations stimulate TOP2B activity within strongly transcribed regions and enhancers. Highly expressed genes that are devoid of usually associated chromatin marks, such as H3K36me3, are deficient in TOP2B activity, indicating that TOP2B may be inefficient at sensing transcription-generated torsional stress directly. Active promoters and the transcription start sites (TSS) with high RNA polymerase II (RNAPII) occupancy show elevated TOP2B ChIP-seq signals but are depleted in TOP2Bccs, indicating that TOP2B is held inactive at active promoters. Surprisingly, TOP2B inhibition with etoposide increased nascent transcription at highly expressed genes and enhancers. While ETP treatment reduced nascent transcription within long genes, these effects were independent of transcript length and instead correlated with the presence of intragenic enhancers. Together these results provide insights into the epigenetic regulation of topoisomerase activity and reveal new roles for TOP2B in neuronal transcriptional regulation. Re-analysis of GSM1516578: TOP2B ChIP-seq in Control with mm10 alignments - Raw FASTQ sequencing reads were analyzed for quality control using FastQC, trimmed using Trimmomatic, and aligned to the mm10 reference genome (GRCm38.p6 assembly) using BWA-MEM. BAM files were marked and filtered for duplicate reads using Sambamba and Samtools, respectively. Peaks were called using MACS2 from biological duplicates and pseudo-replicates where each ChIP sample was normalized to input DNA. MACS2-generated bedGraph files were converted to bigWig files using Kent Utils for individual replicates and pooled replicates