Journal Article

Massively multiplex single-molecule oligonucleosome footprinting

Underwood, Jason G, Goodarzi, Hani, Estes, Laurel S, Narlikar, Geeta J, Ramani, Vijay, Keith, Aidan, Kasinathan, Sivakanthan, Abdulhay, Nour J, Karimzadeh, Mehran, McNally, Colin P, Hsieh, Laura J

2020

Overview

Our understanding of the beads-on-a-string arrangement of nucleosomes has been built largely on high-resolution sequence-agnostic imaging methods and sequence-resolved bulk biochemical techniques. To bridge the divide between these approaches, we present the single-molecule adenine methylated oligonucleosome sequencing assay (SAMOSA). SAMOSA is a high-throughput single-molecule sequencing method that combines adenine methyltransferase footprinting and single-molecule real-time DNA sequencing to natively and nondestructively measure nucleosome positions on individual chromatin fibres. SAMOSA data allows unbiased classification of single-molecular 'states' of nucleosome occupancy on individual chromatin fibres. We leverage this to estimate nucleosome regularity and spacing on single chromatin fibres genome-wide, at predicted transcription factor binding motifs, and across human epigenomic domains. Our analyses suggest that chromatin is comprised of both regular and irregular single-molecular oligonucleosome patterns that differ subtly in their relative abundance across epigenomic domains. This irregularity is particularly striking in constitutive heterochromatin, which has typically been viewed as a conformationally static entity. Our proof-of-concept study provides a powerful new methodology for studying nucleosome organization at a previously intractable resolution and offers up new avenues for modeling and visualizing higher order chromatin structure.

• Publisher: eLife Science Publications, Ltd,eLife Sciences Publications, Ltd,eLife Sciences Publications Ltd
• Subject: Acetylation / Binding Sites / Chromatin / Chromatin - chemistry / Chromatin - genetics / Chromatin - metabolism / Chromosomes and Gene Expression / DNA - chemistry / DNA - genetics / DNA - metabolism / DNA sequencing / Epigenesis, Genetic / Genetics and Genomics / Genomics / High-Throughput Nucleotide Sequencing / high-throughput sequencing / Histones - chemistry / Histones - genetics / Histones - metabolism / Humans / K562 Cells / Nucleic Acid Conformation / nucleosomes / Nucleosomes - chemistry / Nucleosomes - genetics / Nucleosomes - metabolism / Proof of Concept Study / Protein Conformation / Protein Processing, Post-Translational / Single Molecule Imaging / Site-Specific DNA-Methyltransferase (Adenine-Specific) - metabolism / Tools and Resources / Transcription (Genetics) / Transcription Factors - genetics / Transcription Factors - metabolism / Transferases