Étude structurale du complexe formé par la protéine Werner (WRN) et l'hétérodimère Ku70/80

Overview & Synopsis

Chemotherapy or radiobiology treatments aim at generating DNA double-strand breaks (DSBs).Tumor cells are more sensitive to DSBs than healthy cells due their phenotype and genotype. One important axis in radiobiology is to combine radiation therapy with inhibitors of the DNA repair pathways to increase radio-sensitivity and overcome radiation resistance of some cancer cells. Our objectives is to unveil the molecular mechanism of the NHEJ (Non Homologous End Joining) pathwayand to characterize new specific inhibitors of this pathway. Ku70/Ku80 heterodimer (Ku) can recruit several factors of the NHEJ pathway through direct interactions and thus acts as a hub that coordinates the whole NHEJ. Many interactions involve motifscalled KBM (Ku Binding Motif). When DSB ends need to be processed, enzymes arenecessary to correct the extremities prior to the final ligation step. One of the enzymes is Werner (WRN) that has two KBM motifs. During my PhD, I successfully produced several WRN constructs and full-length protein and studied them by combining multiple structural approaches. I showed that WRNcan adopt several oligomeric state using SAXS and SEC-MALS, and that WRN interacts directly with Kuthrough two peptidic motifs, located in N-terminal and C-terminal extremities. I determined the CryoEM structure of a Ku-DNA complex bound to the exonuclease domain of WRN at 3Å of resolution.We showed that Ku recruits WRN through its KBM motif and position WRN near the DNA.