The fate of DNA Double-strand-breaks in plants

DNA double-strand break (DSB) repair is a complex process including DSB formation, processing of broken ends and repair. The fidelity of the reaction determines whether insertion-deletions (indels) are formed upon repair or whether the original DNA sequence is restored. DSBs repair failure can also lead to harmful chromosomal aberrations. Here we describe the fates of DNA DSBs in somatic tissues of plants. We analyzed hundreds of CRISPR-Cas targets and found a great diversity for editing efficiencies via NHEJ as measured by indel formation. A detailed repair kinetics analysis including characterization of repair intermediates at a few sites shows that editing efficiency can be affected by the rate of DSB induction and by precise versus error-prone repair. Repair via homologous recombination (HR) is considered as high-fidelity compared to NHEJ. Using different assays, in tomato and Arabidopsis, we show that gene conversion, one of the outcomes of HR-mediated repair, can account for up to 14% of the repair events. Crossover was a less frequent repair outcome. In addition, we could detect catastrophic events triggered by un-repaired DSBs such as chromosome loss, duplications and inversions that could be explained by induction of a breakage-fusion-bridge cycle, and reshuffling of large chromosomal segments that could be explained by chromothripsis. We discuss the genetic and epigenetic factors that might affect the DSB induction and repair process.