KAS-ATAC reveals the genome-wide single-stranded accessible chromatin landscape of the human genome

Gene regulation in most eukaryotes involves two fundamental processes: alterations in genome packaging by nucleosomes, with active cis-regulatory elements (CREs) generally characterized by open-chromatin configuration, and transcriptional activation. Mapping these physical properties and biochemical activities, through profiling chromatin accessibility and active transcription, is a key tool for understanding the logic and mechanisms of transcription and its regulation. However, the relationship between these two states has not been accessible to simultaneous measurement. To this end, we developed KAS-ATAC, a combination of the kethoxal-assisted ssDNA sequencing (KAS-seq) and assay for transposase-accessible chromatin using sequencing (ATAC-seq) methods for mapping single-stranded DNA (and thus active transcription) and chromatin accessibility, respectively, enabling the genome-wide identification of DNA fragments that are simultaneously accessible and contain ssDNA. We use KAS-ATAC to evaluate levels of active transcription over different CRE classes, to estimate absolute levels of transcribed accessible DNA over CREs, to map nucleosomal configurations associated with RNA polymerase activities, and to assess transcription factor association with transcribed DNA through transcription factor binding site (TFBS) footprinting. We observe lower levels of transcription over distal enhancers compared with promoters and distinct nucleosomal configurations around transcription initiation sites associated with active transcription. We find that most TFs associate equally with transcribed and nontranscribed DNA, but a few factors specifically do not exhibit footprints over ssDNA-containing fragments. We anticipate KAS-ATAC to continue to derive useful insights into chromatin organization and transcriptional regulation in other contexts in the future.