RT Journal Article
SR Electronic
T1 Synthesis of a eukaryotic chromosome reveals a role for N6-methyladenine in nucleosome organization
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 184929
DO 10.1101/184929
A1 Leslie Y. Beh
A1 Galia T. Debelouchina
A1 Kelsi A. Lindblad
A1 Katarzyna Kulej
A1 Elizabeth R. Hutton
A1 John R. Bracht
A1 Robert P. Sebra
A1 Benjamin A. Garcia
A1 Tom W. Muir
A1 Laura F. Landweber
YR 2017
UL http://biorxiv.org/content/early/2017/09/06/184929.abstract
AB Biochemical studies of chromatin have typically used either artificial DNA templates with unnaturally high affinity for histones, or small genomic DNA fragments deprived of their cognate physical environment. It has thus been difficult to dissect chromatin structure and function within fully native DNA substrates. Here, we circumvent these limitations by exploiting the minimalist genome of the eukaryote Oxytricha trifallax, whose notably small ~3kb chromosomes mainly encode single genes. Guided by high-resolution epigenomic maps of nucleosome organization, transcription, and DNA N6-methyladenine (m6dA) locations, we reconstruct full-length Oxytricha chromosomes in vitro and use these synthetic facsimiles to dissect the influence of m6dA and histone post-translational modifications on nucleosome organization. We show that m6dA directly disfavors nucleosomes in a quantitative manner, leading to local decreases in nucleosome occupancy that are synergistic with histone acetylation. The effect of m6dA can be partially reversed by the action of an ATP-dependent chromatin remodeler. Furthermore, erasing m6dA marks from Oxytricha chromosomes leads to proportional increases in nucleosome occupancy across the genome. This work showcases Oxytricha chromosomes as powerful yet practical models for studying eukaryotic chromatin and transcription in the context of biologically relevant DNA substrates.HighlightsDe novo synthesis of complete, epigenetically defined Oxytricha chromosomesEpigenomic profiles of chromatin organization in Oxytricha’s miniature chromosomesm6dA directly disfavors nucleosome occupancy in natural and synthetic chromosomesHistone acetylation and chromatin remodelers temper the impact of m6dA on chromatin