Cicero Predicts cis-Regulatory DNA Interactions from Single-Cell Chromatin Accessibility Data

Linking regulatory DNA elements to their target genes, which may be located hundreds of kilobases away, remains challenging. Here, we introduce Cicero, an algorithm that identifies co-accessible pairs of DNA elements using single-cell chromatin accessibility data and so connects regulatory elements to their putative target genes. We apply Cicero to investigate how dynamically accessible elements orchestrate gene regulation in differentiating myoblasts. Groups of Cicero-linked regulatory elements meet criteria of “chromatin hubs”—they are enriched for physical proximity, interact with a common set of transcription factors, and undergo coordinated changes in histone marks that are predictive of changes in gene expression. Pseudotemporal analysis revealed that most DNA elements remain in chromatin hubs throughout differentiation. A subset of elements bound by MYOD1 in myoblasts exhibit early opening in a PBX1- and MEIS1-dependent manner. Our strategy can be applied to dissect the architecture, sequence determinants, and mechanisms of cis-regulation on a genome-wide scale. sci-ATAC-seq data was collected on human skeletal muscle myoblasts (HSMM) in culture at four timepoints after serum switch to induce differentiation into myotubes, 0 hours, 24 hours, 48 hours and 72 hours. Libraries pooled for sequencing (Experiment 1). An additional experiment was collected using the same system, at 0 hours and 72 hours after serum switch (Experiment 2). Bulk ATAC-seq data was also collected for each of the four timepoints. In addition, sci-ATAC-seq data was collected on an artificial mixture of GM12878 and HL60 cells. Lastly, bulk ATAC-seq data was collected at day 0 and day 7 after serum switch in 54-1 immortalized myoblasts that were transduced with lentivirus carrying small guide RNAs targeting Pbx1, Meis1 or non-targeting controls using lentiCRISPRv2-blast. Cells were allowed time for editing post transduction before differentiation. See publication for details.