SUMMARY
Nuclear processes depend on the organization of chromatin, whose basic units are cylinder-shaped complexes called nucleosomes. A subset of mammalian nucleosomes in situ (inside cells) resembles the canonical structure determined in vitro 25 years ago. Nucleosome structure in situ is otherwise poorly understood. Using cryo-ET and 3-D classification analysis of yeast cells, here we find that canonical nucleosomes account for less than 10% of total nucleosomes expected in situ. In a strain in which H2A-GFP is the sole source of histone H2A, class averages that resemble canonical nucleosomes both with and without an extra density are found ex vivo, but not in situ. These data suggest that the yeast intranuclear environment favors multiple non-canonical nucleosome conformations. Using the structural observations here and the results of previous genomics and biochemical studies, we propose a model in which the average yeast nucleosome’s DNA is partially detached in situ.
Competing Interest Statement
The authors have declared no competing interest.
Footnotes
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The manuscript has undergone multiple rounds of revision, resulting in a nearly complete rewrite. Volta phase plate data has been collected and analyzed for all lamella samples. Only lamellae thinner than 160 nm were used. New histone strains have been created. More controls have been done. More explanations of the nature of cryo-ET artifacts and how to interpret in situ cryo-ET data have been added. A small number of canonical nucleosomes is now detected in situ. The main conclusion is the same.
