Allelic topological centering by transcription factors drives oncogenic multi-enhancer transcriptional regulation [ATAC-seq]

Multiple enhancers, often located across vast genomic distances, regulate key genes in cancer. However, how organization of chromatin topology at individual allele enables cancer type-restricted multi-enhancer gene regulation remains unclear. Using acute protein degradation and time-course population-average chromatin conformation capture in lymphoma, we found that the B-cell lineage-determining transcription factor EBF1 preferentially positions multiple enhancers at loci containing sparsely distributed genes essential for B-cell identity and oncogenesis. Our time-resolved sub-diffraction optical tracing of chromatin architecture of over 100,000 alleles further revealed diverse topological conformations that facilitate multi-enhancer interactions in individual lymphoma cells. Mechanistically, we found that positioning of enhancers at the topological center of the allele is required for their interactions with the target promoter, with EBF1 serving as a barrier to the loop extruding cohesin on the enhancers. These findings, which we demonstrate their generalizability to the T-cell lineage-determining transcription factor TCF1 in T-cell leukemia, suggest that lineage-determining transcription factors establish local radial positioning of enhancers and promoters to enable multi-enhancer regulation of key genes in cancer.