Molecular Cell
Volume 43, Issue 3, 5 August 2011, Pages 369-380
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Article
Nonhistone Scm3 Binds to AT-Rich DNA to Organize Atypical Centromeric Nucleosome of Budding Yeast

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Summary

The molecular architecture of centromere-specific nucleosomes containing histone variant CenH3 is controversial. We have biochemically reconstituted two distinct populations of nucleosomes containing Saccharomyces cerevisiae CenH3 (Cse4). Reconstitution of octameric nucleosomes containing histones Cse4/H4/H2A/H2B is robust on noncentromere DNA, but inefficient on AT-rich centromere DNA. However, nonhistone Scm3, which is required for Cse4 deposition in vivo, facilitates in vitro reconstitution of Cse4/H4/Scm3 complexes on AT-rich centromere sequences. Scm3 has a nonspecific DNA binding domain that shows preference for AT-rich DNA and a histone chaperone domain that promotes specific loading of Cse4/H4. In live cells, Scm3-GFP is enriched at centromeres in all cell cycle phases. Chromatin immunoprecipitation confirms that Scm3 occupies centromere DNA throughout the cell cycle, even when Cse4 and H4 are temporarily dislodged in S phase. These findings suggest a model in which centromere-bound Scm3 aids recruitment of Cse4/H4 to assemble and maintain an H2A/H2B-deficient centromeric nucleosome.

Highlights

► Reconstitution of two distinct populations of Cse4 nucleosomes ► Octameric nucleosomes containing Cse4/H4/H2A/H2B favored on noncentromere DNA ► Atypical nucleosomes containing Cse4/H4/Scm3 favored on AT-rich centromere DNA ► Scm3 binds to AT-rich DNA in vitro, enriched at centromeres throughout cell cycle

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