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

Abstract 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.