Topology-informed regulatory element collections coordinate cell identity gene expression programs

Cis-regulatory elements (CREs) contact target genes to regulate their transcription, but CRE interactions are complex and predicting their function is challenging. Contemporary approaches to predict CRE targets generally assume one-to-one associations and linear genomic proximity. These methods fail to account for long-range CRE contacts caused by folding of the genome, for circumstances in which a single CRE regulates multiple genes, or, conversely, when a single gene is regulated by multiple CREs. To address these challenges and investigate three-dimensional (3D) CRE communities, we developed BOUQUET, an integrative, graph-theory-based approach that uses multiple aspects of genome topology to probe communities of CREs, their bound apparatus, and their target genes. Using this approach, we uncover communities of CREs that are undetectable by existing unbiased methods, such as those with interactions that span insulating loop boundaries, and regulatory communities that comprise clustered promoter elements. A subset of these communities, which we term "3D super-enhancer CRE communities" (3DSE CRE communities), accrues exceptional amounts of regulatory apparatus at cell identity-defining genes and evolves the super-enhancer concept. In imaging assays, we observe co-localization and co-expression of in-community genes within puncta of transcriptional co-factors, consistent with microcompartment biology. Genome-wide, pairs of genes within the same 3D CRE community are co-expressed in single cells, and pairs of CREs are co- accessible, suggesting both chromatin activity and transcription within a topologically defined community is coordinated. Our model more accurately reflects the complexity of CRE interaction networks, underscores the need for approaches that incorporate spatial organization in the study of enhancer biology, and provides a framework for the reinterpretation of SEs, their condensates, and their target genes through the lens of 3D chromatin structure. ### Competing Interest Statement The authors have declared no competing interest.