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PMID: 10654939 Published · ppublish English Journal Article Research Support, U.S. Gov't, P.H.S.

The organized chromatin domain of the repressed yeast a cell-specific gene STE6 contains two molecules of the corepressor Tup1p per nucleosome.

The EMBO journal ·Vol. 19 ·No. 3 ·2000-02-01 ·Pages 400-9

Ducker CE, Simpson RT

Abstract

In yeast alpha cells the a cell-specific genes STE6 and BAR1 are packaged as gene-sized chromatin domains of positioned nucleosomes. Organized chromatin depends on Tup1p, a corepressor that interacts with the N-terminal regions of H3 and H4. If Tup1p functions to organize or stabilize a chromatin domain, the protein might be expected to be present at a level stoichiometric with nucleosomes. Chromatin immunoprecipitation assays using Tup1p antibodies showed Tup1p to be associated with the entire genomic STE6 coding region. To determine stoichiometry of Tup1p associated with the gene, a yeast plasmid containing varying lengths of the STE6 gene including flanking control regions and an Escherichia coli lac operator sequence was constructed. After assembly into chromatin in vivo in Saccharomyces cerevisiae, minichromosomes were isolated using an immobilized lac repressor. In these experiments, Tup1p was found to be specifically associated with repressed STE6 chromatin in vivo at a ratio of about two molecules of the corepressor per nucleosome. These observations strongly suggest a structural role for Tup1p in repression and constrain models for organized chromatin in repressive domains.

MeSH Terms
Chromatin/genetics,metabolism DNA, Fungal/genetics Escherichia coli Fungal Proteins/genetics,metabolism Lac Operon Nuclear Proteins Nucleosomes/genetics Plasmids/genetics Precipitin Tests Repressor Proteins/genetics,metabolism Saccharomyces cerevisiae/genetics Saccharomyces cerevisiae Proteins
Chemicals
Chromatin DNA, Fungal Fungal Proteins Nuclear Proteins Nucleosomes Repressor Proteins Saccharomyces cerevisiae Proteins TUP1 protein, S cerevisiae
Authors & Affiliations
2 authors, click to expand affiliations / ORCID
Ducker C E
Department of Biochemistry and Molecular Biology, 308 Althouse, Pennsylvania State University, University Park, PA 16802, USA.
Simpson R T
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Article Info
Journal
The EMBO journal
Abbr.
EMBO J
ISSN
0261-4189
DOI
10.1093/emboj/19.3.400
Published
2000-02-01
Pages
400-9
Language
English
Region
England
NLM ID
8208664
PMCID
PMC305577
Subset
IM
Grants
NIGMS NIH HHS · R01 GM052311 · United States
NIGMS NIH HHS · GM52311 · United States
External Links
PubMed source DOI: 10.1093/emboj/19.3.400 PMC Full Text View journal details
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