The Interplay Between Arginine Dimethylation and Ubiquitylation Coordinates TDP1 Proteostasis for the Repair of Topoisomerase I Covalent Complexes

Abstract

Tyrosyl-DNA phosphodiesterase (TDP1) hydrolyzes the phosphodiester bond between a DNA 3'-end and a tyrosyl moiety and is implicated in the repair of trapped topoisomerase I (Top1)-DNA covalent complexes (Top1cc). Protein arginine methyltransferase 5 (PRMT5) catalyzes methylation of TDP1 at residues R361 and R586. Here we establish mechanistic crosstalk between TDP1 arginine methylation and ubiquitylation, which is critical for TDP1 homeostasis and responses to Top1 poisons. Using PRMT5 knockout cells, we show that loss of TDP1 arginine methylation results in an imbalance in TDP1 degradation with marked elevation of Top1cc-induced DNA breaks and cell death. In the absence of Camptothecin (CPT), we identify that R586 methylation promotes TDP1 ubiquitylation facilitating ubiquitin/proteasome-dependent TDP1 turnover. Abrogation of TDP1 methylation at R586 promotes increased binding with the deubiquitylase UCHL3 and suppresses TDP1 ubiquitination and turnover. On the contrary, CPT-induced DNA damage represses TDP1 ubiquitination and promotes TDP1-XRCC1 binding and formation of XRCC1 repair foci at Top1cc-induced DNA damage sites. We further show R361 dimethylation enhances the 3'-phosphodiesterase activity of TDP1 in real-time fluorescence-based cleavage assays and was rationalized by using structural modeling. Together, our findings establish arginine methylation as a coregulator of TDP1 proteostasis and activity which modulates repair of trapped Top1cc.