Accelerated chromatin biochemistry using DNA-barcoded nucleosome libraries

• Published in: Nature methods Vol. 11; no. 8; pp. 834 - 840
• Main Authors: Nguyen, Uyen T T, Bittova, Lenka, Müller, Manuel M, Fierz, Beat, David, Yael, Houck-Loomis, Brian, Feng, Vanessa, Dann, Geoffrey P, Muir, Tom W
• Format: Journal Article
• Language: English
• Published: New York Nature Publishing Group US 01.08.2014; Nature Publishing Group
• Subjects: 13/106; 45/77; 631/1647/2210/2211; 631/45/612/100; 631/45/612/822; 631/92/96; 82; 82/1; 82/47; 82/58; 82/75; 82/79; 82/80; 82/83; Acetylation; Biochemistry; Bioinformatics; Biological Microscopy; Biological Techniques; Biomedical Engineering/Biotechnology; Chromatin; Chromatin - chemistry; Chromatin Immunoprecipitation; Deoxyribonucleic acid; DNA; DNA Barcoding, Taxonomic; DNA sequencing; Histones; Life Sciences; Methods; Nucleosomes; Nucleosomes - chemistry; Nucleotide sequencing; Observations; Properties; Proteomics
• ISSN: 1548-7091; 1548-7105
• DOI: 10.1038/nmeth.3022

A synthetic library of nucleosomes, each with a DNA-barcode characteristic for a defined post-translational modification (PTM), is used to probe PTM-based recruitment and modulation of histone mark readers and writers. Elucidating the molecular details of how chromatin-associated factors deposit, remove and recognize histone post-translational modification (PTM) signatures remains a daunting task in the epigenetics field. We introduce a versatile platform that greatly accelerates biochemical investigations into chromatin recognition and signaling. This technology is based on the streamlined semisynthesis of DNA-barcoded nucleosome libraries with distinct combinations of PTMs. Chromatin immunoprecipitation of these libraries, once they have been treated with purified chromatin effectors or the combined chromatin recognizing and modifying activities of the nuclear proteome, is followed by multiplexed DNA-barcode sequencing. This ultrasensitive workflow allowed us to collect thousands of biochemical data points revealing the binding preferences of various nuclear factors for PTM patterns and how preexisting PTMs, alone or synergistically, affect further PTM deposition via cross-talk mechanisms. We anticipate that the high throughput and sensitivity of the technology will help accelerate the decryption of the diverse molecular controls that operate at the level of chromatin.