We trapped catalytically engaged topoisomerase IIβ (TOP2B) in covalent DNA cleavage complexes (TOP2Bccs) and mapped their positions genome-wide in cultured mouse cortical neurons. We report that TOP2Bcc distribution varies with both nucleosome and compartmental chromosome organization. While TOP2Bccs in gene bodies correlate with their level of transcription, highly expressed genes that lack the usually associated chromatin marks, such as H3K36me3, show reduced TOP2Bccs, suggesting that histone posttranslational modifications regulate TOP2B activity. Promoters with high RNA polymerase II occupancy show elevated TOP2B chromatin immunoprecipitation sequencing signals but low TOP2Bccs, indicating that TOP2B catalytic engagement is curtailed at active promoters. Surprisingly, either poisoning or inhibiting TOP2B increases nascent transcription at most genes and enhancers but reduces transcription within long genes. These effects are independent of transcript length and instead correlate with the presence of intragenic enhancers. Together, these results clarify how cells modulate the catalytic engagement of topoisomerases to affect transcription.