Reshuffling yeast chromosomes with CRISPR/Cas9

Author summary Chromosomes are highly dynamic objects that often undergo large structural variations such as reciprocal translocations. Such rearrangements can have dramatic functional consequences, as they can disrupt genes, change their regulation or create novel fusion genes at their breakpoints. For instance, 90–95% of patients diagnosed with chronic myeloid leukemia carry the Philadelphia chromosome characterized by a reciprocal translocation between chromosomes 9 and 22. In addition, translocations reorganize the genetic information along chromosomes, which in turn can modify the 3D architecture of the genome and potentially affect its functioning. Quantifying the fitness impact of translocations independently from the confounding effect of base substitutions has so far remained challenging. Here, we report a novel CRISPR/Cas9-based technology allowing to generate with high efficiency and at a base-pair precision either uniquely targeted or multiple reciprocal translocations in yeast, without leaving any marker or scar in the genome. Engineering targeted reciprocal translocations allowed us for the first time to untangle the phenotypic impacts of large chromosomal rearrangements from that of point mutations. In addition, the generation of multiple translocations led to a large reorganization of the genetic information along the chromosomes, often including unanticipated large segmental duplications. We showed that reshuffling the genome resulted in the emergence of fitness advantage in stressful environmental conditions, even in strains where no gene was disrupted or amplified by the translocations.