LAUSR

Summary Polyubiquitination of proliferating cell nuclear antigen (PCNA) regulates the error-free template-switching mechanism for the bypass of DNA lesions during DNA replication. PCNA polyubiquitination is critical for the maintenance of genomic integrity; however, the underlying mechanism is poorly understood. Here, we demonstrate that tonicity-responsive enhancer-binding protein (TonEBP) regulates PCNA polyubiquitination in response to DNA damage. TonEBP was recruited to DNA damage sites with bulky adducts and sequentially recruited E3 ubiquitin ligase SHPRH, followed by deubiquitinase USP1, to DNA damage sites, in correlation with the dynamics of PCNA polyubiquitination. Similarly, TonEBP was found to be required for replication fork protection in response to DNA damage. The Rel-homology domain of TonEBP, which encircles DNA, was essential for the interaction with SHPRH and USP1, PCNA polyubiquitination, and cell survival after DNA damage. The present findings suggest that TonEBP is an upstream regulator of PCNA polyubiquitination and of the DNA damage bypass pathway.