Abstract B010: DNA-PKcs-dependent phosphorylation of RECQL4 promotes NHEJ by stabilizing the NHEJ machinery at DNA double-strand breaks

Abstract

Abstract RECQL4, one of the five RecQ helicases in mammalian cells, guards the genome by promoting DNA replication, DNA repair, and telomere maintenance. Mutations in human RECQL4 are associated with three genetic disorders, Rothmund-Thomson syndrome (RTS), Baller-Gerold syndrome (BGS), and RAPADILINO syndrome, which display accelerated aging, developmental abnormalities, and a predisposition to cancer. It is believed that accumulation of DNA damage contributes to the clinical features observed in patients with RECQL4 mutations. Previously, RECQL4 was implicated in promoting the DNA double strand break repair pathway non-homologous end joining (NHEJ), but its role in the pathway remains unresolved. In this study, we report that RECQL4 stabilizes the NHEJ machinery at DSBs to promote repair. Specifically, we find that RECQL4 interacts with the NHEJ core factor DNA-PKcs and the interaction is increased following IR. RECQL4 promotes DNA end bridging mediated by DNA-PKcs and Ku70/80 in vitro and the accumulation/retention of NHEJ factors at DSBs in vivo. Moreover, interaction between DNA-PKcs and the other core NHEJ proteins following IR treatment is attenuated in the absence of RECQL4. These data indicate that RECQL4 promotes the stabilization of the NHEJ factors at DSBs to support formation of the NHEJ long-range synaptic complex. In addition, we observed that the kinase activity of DNA-PKcs is required for accumulation of RECQL4 to DSBs and that DNA-PKcs phosphorylates RECQL4 at six serine/threonine residues. Blocking phosphorylation at these sites reduced the recruitment of RECQL4 to DSBs, attenuated the interaction between RECQL4 and NHEJ factors, destabilized interactions between the NHEJ machinery, and resulted in decreased NHEJ. Collectively, these data illustrate reciprocal regulation between RECQL4 and DNA-PKcs in NHEJ and the potential that dysfunctional NHEJ may play a role in RECQL4-associated genetic disorders and tumorigenesis. Citation Format: Anthony J. Davis. DNA-PKcs-dependent phosphorylation of RECQL4 promotes NHEJ by stabilizing the NHEJ machinery at DNA double-strand breaks [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Damage Repair: From Basic Science to Future Clinical Application; 2024 Jan 9-11; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2024;84(1 Suppl):Abstract nr B010.