Assignment of structural transitions during mechanical unwrapping of nucleosomes and their disassembly products

Abstract

Nucleosome DNA unwrapping and its disassembly into hexasomes and tetrasomes is necessary for genomic access and plays an important role in transcription regulation. Previous single-molecule mechanical nucleosome unwrapping revealed a low- and a high-force transitions, and force-FRET pulling experiments showed that DNA unwrapping is asymmetrical occurring always first from one side before the other. However, the assignment of DNA segments involved in these transitions remains controversial. Here, using high-resolution optical tweezers with simultaneous single-molecule FRET detection we show that the low-force transition corresponds to the undoing of the outer-wrap of one side of the nucleosome (~27 bp), a process that can occur either cooperatively or non-cooperatively, whereas the high-force transition corresponds to the simultaneous unwrapping of ~76 bp from both sides. This process may give rise stochastically to the disassembly of nucleosomes into hexasomes and tetrasomes whose unwrapping/rewrapping trajectories we establish. In contrast, nucleosome rewrapping does not exhibit asymmetry. To rationalize all previous nucleosome unwrapping experiments, it is necessary to invoke that mechanical unwrapping involves two nucleosome reorientations: one that contributes to the change in extension at the low-force transition, and another that coincides but does not contribute to the high-force transition.

Assignment of structural transitions during mechanical unwrapping of nucleosomes and their disassembly products

“…The change in extension analysis code and the mechanical unwrapping model code have been deposited in GitHub ( https://github.com/CesarDiazCelis1/Nucleosome-Unwrapping ) ( 59 ). The optical tweezers data are available via Zenodo ( https://doi.org/10.5281/zenodo.6780532 ) ( 60 ).…”