Abstract
We demonstrate that the yeast flocculation gene, FLO1, is representative of a distinct subset of subtelomeric genes that are robustly repressed by the Cyc8–Tup1 complex. We have examined Cyc8–Tup1 localisation, histone acetylation and long-range chromatin remodelling within the extensive FLO1 upstream region. We show that Cyc8–Tup1 is localised in a DNase I hypersensitive site within an ordered array of strongly positioned nucleosomes around −700 base pairs upstream of the transcription start site. In cyc8 deletion mutant strains, Tup1p localisation is absent, with concomitant histone hyperacetylation of adjacent regions at the FLO1 promoter. This is accompanied by extensive histone depletion across the upstream region and gene activation. The yeast histone deacetylases, Hda1p and Rpd3p, occupy the repressed FLO1 promoter region in a Cyc8–Tup1 dependent manner and coordinate histone deacetylation, nucleosome stabilisation and gene repression. Moreover, we show that the ATP-dependent chromatin remodelling complex Swi–Snf occupies the site vacated by Cyc8–Tup1 in a cyc8 mutant. These data suggest that distinctly bound Cyc8–Tup1 cooperates with Hda1p and Rpd3p to establish or maintain an extensive array of strongly positioned, deacetylated nucleosomes over the FLO1 promoter and upstream region which inhibit histone acetylation, block Swi–Snf binding and prevent transcription.
Highlights
Co-repressors and co-activators play a key role in eukaryotic gene regulation
We confirmed that Cyc8–Tup1 binds to a distinct site at the FLO1 promoter where it is associated with an array of strongly positioned, deacetylated nucleosomes covering the promoter and upstream region
We revealed that in the absence of Cyc8–Tup1, there is a gross remodelling of FLO1 promoter and upstream chromatin involving nucleosome acetylation, rearrangement and eviction which accompanies FLO1 gene de-repression
Summary
Co-repressors and co-activators play a key role in eukaryotic gene regulation. Through association with DNA-bound transcription factors in the nucleus, these often large complexes determine the repressed or active state of gene promoters by altering the chromatin structure or the recruitment and competence of the RNA polymerase II holoenzyme. One of the earliest co-repressors of gene regulation identified in the yeast, Saccharomyces cerevisiae, was the Cyc8–Tup complex; a 1.4 megadalton complex composed of the TUP1 and CYC8 ( known as SSN6) gene products in a 4:1 ratio [1,2]. The complex is formed by the interaction of the N-terminal residues of Tup1p with tetratricopeptide repeat (TPR) motifs of Cyc8p [3,4,5]. Tup1p is related to the Groucho family of co-repressor proteins in higher eukaryotes by way of its C-terminal WD-repeat domain [6].
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