ATAD5 Dual-Domain Interaction with UAF1 Enhances USP1-mediated PCNA De-ubiquitination in the TLS Pathway

Translesion synthesis (TLS) bypasses DNA lesions by using error-prone DNA polymerases recruited by mono-ubiquitinated PCNA (Ub-PCNA). To reduce the mutagenic consequences of TLS, the USP1/UAF1 complex removes ubiquitin from PCNA, terminating the TLS process. This study investigated the role of ATAD5 in the PCNA de-ubiquitination (DUB). We demonstrated that ATAD5 binds USP1/UAF1 and Ub-PCNA simultaneously as a molecular adaptor, connecting the DUB enzyme and the substrate to promote DUB reactions. The SIM domain of ATAD5 binds the SUMO-like domains (SLD1 and SLD2) of UAF1 and has canonical interactions with SLD2. Furthermore, we found that the disordered linker of ATAD5 contributes partially to its regulatory activity. Our structural studies of UAF1/SIM chimeras yielded crystals diffracting to 7.6 Å resolution for SIM-SLD interface analysis. These findings provide new insights into the molecular mechanisms of TLS regulation, underscoring the potential of ATAD5 as a therapeutic target for inducing genomic instability in cancer cells.