Abstract 1419: Role of a novel Senataxin-associated nuclease in DNA repair

Abstract

Abstract Yeast two-hybrid analysis has identified a novel, uncharacterized Senataxin-interacting protein with 5’ exonuclease activity, and homology to the FEN1 nuclease domain. Senataxin is an RNA/DNA helicase that functions in resolving R loop structures which occur as a result of normal replication and transcription but can also persist resulting in genomic instability. We found that SAN1 (Senataxin-Associated Nuclease 1) is unable to cut dsDNA or bubbles, but is active against 5’ overhangs and against ssDNAs of greater than 15 nucleotides. SAN1 cleaves 3 or 8 nt fragments from the 5’ end of ssDNA and utilizes magnesium as a cofactor. As the definitive 5’ nuclease responsible for unhooking ICLs has yet to be identified we speculate that SAN1 might function in excising crosslinked nucleotides that result from interstrand crosslink (ICL) damage. We generated a SAN1 -/- HeLa cell line through CRISPR/Cas gene editing and found that SAN1 -/- cells are sensitized to ICL agents mitomycin-c and cisplatin. Interestingly, colony survival assays have shown that SAN1 does not appear to be epistatic of FANCD2, a key component of the classical pathway for resolving ICLs, the Fanconi Anemia pathway. Additionally, although the nuclease domain of SAN1 is homologous to the FEN1 family of structure-specific nucleases, SAN1 also possesses a unique and conserved C terminus of unknown function. Our goal is to understand the relationship between the structure and nuclease activity of SAN1 in DNA repair and how SAN1 relates to other known ICL-repair pathways. Understanding the components that regulate genome stability is crucial in understanding how cancer occurs and what protein targets will be the most advantageous to pursue in developing potential therapeutics. This study may also provide a better understanding of a novel factor in resistance to ICL-inducing chemotherapeutics such as cisplatin and MMC which are still widely used in the treatment of breast and ovarian cancers. Citation Format: Heather McCartney, Ian Macara. Role of a novel Senataxin-associated nuclease in DNA repair [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1419. doi:10.1158/1538-7445.AM2017-1419