Séminaire de l’Institut Jacques Monod – Amir Aharoni

Invité par l’équipe Palancade, Amir Aharoni (Department of Life Sciences and the National Institute for Biotechnology in the Negev, Ben-Gurion University of the Negev, Israel) présentera un séminaire de l’Institut Jacques Monod sur le thème :

Real-time monitoring of replication fork progression through damaged DNA

Résumé :

Despite the frequent occurrence of AP sites in the eukaryotic genome, understanding how replication forks bypass non-repaired AP sites remains elusive. While an error-prone and an error-free DNA damage bypass pathways exist, the choice and timing of activation of each bypass pathway during AP sites replication are still unknown. Here, we have established and examined a system for measuring replication fork progression through site-specific AP sites in live yeast cells, for monitoring DNA damage bypass in real time. In this system, AP sites are introduced site-specifically between two bacterial fluorescent repressor-operator systems (FROS) during the G1 phase, using catalytically inactive Cas9 fused to DNA glycosidase. Time-lapse fluorescent microscopy is then used to monitor the progression of single replisomes through the AP sites and to reveal the timing of AP bypass. Using this real-time microscopy system, we revealed the cellular choice between replication associated and post replicative AP sites bypass pathways controlling replication fork stalling at AP sites and cell survival.