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Biochem Biophys Res Commun. 2017 Jan 22;482(4):896-901. doi: 10.1016/j.bbrc.2016.11.130. Epub 2016 Nov 24.

In vitro reconstitution and biochemical analyses of the Schizosaccharomyces pombe nucleosome.

Author information

1
Laboratory of Structural Biology, Graduate School of Advanced Science and Engineering, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan.
2
Advanced ICT Research Institute Kobe, National Institute of Information and Communications Technology, 588-2 Iwaoka, Iwaoka-cho, Nishi-ku, Kobe 651-2492, Japan.
3
Advanced ICT Research Institute Kobe, National Institute of Information and Communications Technology, 588-2 Iwaoka, Iwaoka-cho, Nishi-ku, Kobe 651-2492, Japan; Graduate School of Frontier Biosciences, Osaka University, 1-3 Yamadaoka, Suita, Osaka 565-0871, Japan.
4
Laboratory of Structural Biology, Graduate School of Advanced Science and Engineering, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan; Research Institute for Science and Engineering, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan; Institute for Medical-oriented Structural Biology, Waseda University, 2-2 Wakamatsu-cho, Shinjuku-ku, Tokyo 162-8480, Japan. Electronic address: kurumizaka@waseda.jp.

Abstract

Schizosaccharomyces pombe, which has a small genome but shares many physiological functions with higher eukaryotes, is a useful single-cell, model eukaryotic organism. In particular, many features concerning chromatin structure and dynamics, including heterochromatin, centromeres, telomeres, and DNA replication origins, are well conserved between S. pombe and higher eukaryotes. However, the S. pombe nucleosome, the fundamental structural unit of chromatin, has not been reconstituted in vitro. In the present study, we established the method to purify S. pombe histones H2A, H2B, H3, and H4, and successfully reconstituted the S. pombe nucleosome in vitro. Our thermal stability assay and micrococcal nuclease treatment assay revealed that the S. pombe nucleosome is markedly unstable and its DNA ends are quite accessible, as compared to the canonical human nucleosome. These findings are important to understand the mechanisms of epigenetic genomic DNA regulation in fission yeast.

KEYWORDS:

Histones; Micrococcal nuclease; Nucleosome; Reconstitution; Schizosaccharomyces pombe; Thermal stability

PMID:
27890612
DOI:
10.1016/j.bbrc.2016.11.130
[Indexed for MEDLINE]

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