Large-scale discovery of chromatin dysregulation induced by oncofusions and other protein-coding variants

Although population-scale databases have expanded to millions of genetic variants, mechanistic insight into variant function has not kept pace. We present PROD-ATAC, a high-throughput method for discovering the effects of protein-coding variants on chromatin regulation. A pooled library of variants is expressed in a disease-agnostic biosensor cell line, and single-cell ATAC resolves each variant’s effect on chromatin. Using PROD-ATAC, we characterized the impacts of >100 oncogenic fusions and controls. We showed that pioneer activity is a common feature of fusions spanning an enormous range of frequencies. Further, fusion-induced dysregulation can be context-agnostic as observed mechanisms overlapped with cancer and cell-type specific prior knowledge. We also showed that gain-of-function pioneering is a common feature among oncofusions. This work provides a global view of fusion-induced chromatin disruption across many fusion frequencies and cancer types. Systematically dissecting this landscape revealed convergent mechanisms among seemingly disparate oncofusions and shared modes of chromatin dysregulation across different cancers. A pooled library of >100 oncofusions and controls was expressed in HEK293T cells where each cell contained one protein-coding perturbation. 10X Genomics' droplet platform was used to capture ~120,000 nuclei (~10,000 nuclei per sample for 12 samples) and Spear-ATAC was used to resolve the effects of each variant on chromatin regulation. Note in this case the 12 samples all dervied from the same biological library of cells and are in this way replicates used to increase the number of nuclei captured (but they are not different biological conditions).