Since the advent of chromatin conformation capture (3C) technology, our understanding of the physical organization of the genome has expanded into 3D. These data enabled the discovery of topologically associating domains and chromatin looping and how this spatial packaging of DNA impacts gene regulation. There remains, however, a gap spanning the single nucleotide information captured in linear NGS epigenetic assays such as nucleosome positioning and histone modifications and the 100’s kb to mb looping events that link CTCF anchors through cohesion-rings and enhancer-promoter interactions.
This gap exists due to the technical limitations caused by restriction-enzyme-based digestion, where the molecular question is asked, “where are restriction sites and how are they interacting?” This approach misses out on the fundamental building block of chromatin – the nucleosome. Understanding how nucleosomes influence genomic structure and gene regulation by chromatin remodeling, histone modification, and distal interactions is critical in building regulatory models and unraveling the mechanical processes that drive transcriptional programming. The use of MNase is a relatively new approach to 3C to target the interactome from the viewpoint of the nucleosome. This approach hits a trifecta of requirements for bridging this gap in the chromatin knowledge: 1) Nucleosomes are evenly positioned across the genome, therefore includes more of the genome in the sequence data, 2) Fragments generated by MNase are very consistent in size, providing a stable building block to accumulate sequence data, 3) the size of the fragment is about the length of a sequence length (2×150), meaning each read covers an entire nucleosome-length fragment. These aspects of MNase-based topology provide a high-resolution objective lens to the conformation microscope that is 3C. Whether it’s a protein-directed or genome-wide view, nucleosome-centric topology enables the interrogation and description of chromatin ultrastructure, allowing researchers an unparalleled view into chromatin dynamics.