Abstract 6101: PUM3 is a novel R-loop regulator and is essential for efficient DNA damage repair

Abstract

Abstract R-loops have important roles in regulating transcription and genome integrity. To identify novel proteins that function to modulate DNA damage repair, we screened a library of CRISPR-engineered RNA binding protein knock-out cells. From this analysis, we found that a poorly characterized protein, PUM3, is important DNA damage repair and that PUM3-deficient cells were very sensitive the chemotoxic agents. We show that under normal conditions, PUM3 is localized to the nucleolus and nucleus, where it binds to R-loops and modulates transcription. In this setting, PUM3 functions to suppress/resolve R-loops and that PUM3-/- cells have elevated R-loop levels. Following DNA damage, PUM3 is rapidly re-localized from the nucleolus to sites of DNA damage in the nucleus by members of the DDX/DHX family of RNA-helicases. Using small molecule inhibitors and quantitative assays, we find that PUM3 functions in homologous recombination (HR) mediated DNA repair and that PUM3-/- cells have significantly reduced HR-efficiency. PUM3 function at sites of DNA breaks are not affected by transcription or R-loops, implicating PUM3 as functioning after DNA resection during DNA repair. Analysis of TCGA patient data shows that PUM3 levels correlate with Cisplatin resistance and patient outcome. Collectively, our data shows that PUM3 is a previously unknown regulator of R-loops and DNA damage repair that has an important role in chemotherapy sensitivity in cancer patients. Citation Format: Chenyu Lin, Jian Ouyang, Robert Wine, Chen Qiu, Jason Williams, Traci M. Hall, Lee Zou, Wayne Miles. PUM3 is a novel R-loop regulator and is essential for efficient DNA damage repair [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6101.