Structural and functional insights into the interaction between Ku70/80 and Pol X family polymerases in NHEJ
Philippe Frit, Himani Amin, Sayma Zahid, Nadia Barboule, Chloe M. Hall, Gurdip Matharu, Steven W. Hardwick, Jeanne Chauvat, Sébastien Britton, Dimitri Y. Chirgadze, Virginie Ropars, Jean‐Baptiste Charbonnier, Patrick Calsou, Amanda K. Chaplin
Abstract
Non-homologous end joining (NHEJ) is the main repair pathway for double-strand DNA breaks (DSBs) in mammals. DNA polymerases lambda (Pol λ) and mu (Pol μ), members of the Pol X family, play a key role in this process. However, their interaction within the NHEJ complexes is unclear. Here, we present cryo-EM structures of Pol λ in complex with the DNA-PK long-range synaptic complex, and Pol μ bound to Ku70/80-DNA. These structures identify interaction sites between Ku70/80 and Pol X BRCT domains. Using mutants at the proteins interface in functional assays including cell transfection with an original gap-filling reporter, we define the role of the BRCT domain in the recruitment and activity of the two Pol X members in NHEJ and in their contribution to cell survival following DSBs. Finally, we propose a unified model for the interaction of all Pol X members with Ku70/80.