BioProject

Spatial genome organization is essential to direct fundamental DNA-templated biological processes (e. More...

Spatial genome organization is essential to direct fundamental DNA-templated biological processes (e.g. transcription, replication, and repair), but the 3D in situ nanometer-scale structure of accessible cis-regulatory DNA elements within the crowded nuclear environment remains elusive. Here, we combined the recently developed Assay for Transposase-Accessible Chromatin with visualization (ATAC-see), PALM super-resolution imaging and lattice light-sheet microscope (a method termed 3D ATAC-PALM) to selectively image and quantitatively analyze key features of the 3D accessible genome in single cells. 3D ATAC-PALM reveals that accessible chromatin are non-homogeneously organized into spatially segregated clusters or accessible chromatin domains (ACDs). To directly link imaging and genomic data, we optimized multiplexed imaging of 3D ATAC-PALM with Oligopaint DNA-FISH, RNA-FISH and protein fluorescence. We found that ACDs colocalize with active chromatin and enclose transcribed genes. By applying these methods to analyze genetically purterbed cells, we demonstrated that genome architectural protein CTCF prevents excessive clustering of accessible chromatin and decompacts ACDs. These results highlight the 3D ATAC-PALM as a useful tool to probe the structure and organizing mechanism of the genome. Overall design: Examine accessible chromatin landscape (bulk ATAC-seq) in mouse embryonic stem cells under a variety of chemical/genetic purturbations. We performed at least two biological replicates for each condition. We also performed Micro-C experiment after acute auxin-induced acute CTCF depeltion for 6 hours. Less...