Arginine methylation of human DNA topoisomerase I by PRMT5 facilitates DNA relaxation.

DNA topoisomerase 1 (Top1) is essential for resolving DNA supercoiling during replication and transcription. Here, we identify protein arginine methyltransferase 5 (PRMT5) as a novel regulator of human Top1 activity via symmetric dimethylation at arginine residues R708 and R749, located in the linker and catalytic domains, respectively. Methylation enhances Top1-mediated strand rotation and DNA relaxation without affecting its DNA binding ability. In contrast, methylation-deficient Top1 mutants (Top1KK) display impaired subnuclear mobility and accumulate elevated levels of trapped Top1-DNA covalent complexes (Top1cc) upon camptothecin (CPT) treatment. These defects are independent of PRMT5-Top1 binding but are dependent on PRMT5's enzymatic activity. Loss of Top1 methylation-via point mutation, PRMT5 knockout, or pharmacological inhibition-delays Top1cc resolution and amplifies CPT-induced DNA damage. Strikingly, combining PRMT5 inhibitors (PRMT5i) with Top1 poisons such as irinotecan enhances cytotoxicity across multiple cancer cell types. In a triple-negative breast cancer mouse model, this combination significantly suppresses tumor growth and metastasis, accompanied by increased DNA damage. Our results define PRMT5-driven Top1 arginine methylation as a crucial regulatory mechanism and highlight PRMT5i as a means to potentiate Top1-based cancer treatment.