Spatial Genome Architecture in Development & Disease

Dissecting the role of chromosome genome organization in chromosome fragility and the formation of leukemia driving translocations


Processes that traverse DNA are associated with dramatic changes in DNA topology. These changes occur in the context of the chromatin and must be coordinated with processes that shape chromosome organization per se, such as the formation of chromatin loops and the higher-order chromosome structure. Accumulating topological stress is dissipated at strategic genomic locations, such as promoters and chromatin loop boundaries, by the action of enzymes called topoisomerases. Type II topoisomerases (TOP2), incise both strands of DNA to release supercoiling, unknot, and decatenate DNA. Abortive TOP2 actions induced by the presence of a widely used class of chemotherapeutics, called topoisomerase poisons, however, stabilize TOP2s on DNA, promoting chromosome breakage within recurrent fragile sites that often form chromosome fusions that drive secondary, therapyrelated leukemias. The contribution of cellular processes in triggering the conversion of stabilized TOP2 to DNA breaks that promote the formation of oncogenic translocations remains elusive.  Driven by our preliminary results, we propose that transcription and processes that shape chromosome folding are essential contributors to chromosome fragility that form oncogenic fusions. Our aim, therefore, is to directly assess the effect of the 3D chromosome organization to two different aspects of the formation of recurrent fusions: (1) the role of chromatin loop dynamics to chromosome fragility at recurrent translocation hot spots and (2) the role of spatial genome organization to the chromosome-end synapsis, a crucial step of the translocation process that dictates the choice of partners. We will directly assess the effects of chromatin loop dynamics to TOP2-induced genomic instability by using state-of-the-art next-generation sequencing approaches to profile DNA breaks across the genome, in cellular systems allowing the formation or elimination of chromatin loops, in a controlled fashion. We will also directly evaluate a causal relationship of the 3D nuclear architecture with translocation formation by combining high-throughput imaging methodologies we have developed to probe chromosome breakage, synapsis and fusions, with precision genome editing to perturb gene activity and chromosome structure at single cell resolution. We anticipate that our proposed work will set 3D chromatin folding as a major contributor to the cellular aetiology of recurrent oncogenic fusions.