Figure 1. Histone deacetylation is required for proper telomere structure.

(A) Construct 2 consists of the URA3 gene followed by a downstream Gal UAS with a mutated TATA box (UAS*). When integrated at telomere 7L, the UAS* folds back and drives URA3 transcription in the presence of galactose. (B) Using the SGA (synthetic genetic array) technology, construct 2 was introduced into the ∼4800 strains of the viable haploid yeast gene deletion collection. Subsequently cells were robotically pinned onto −FOA and +FOA galactose-containing media in quadruplicate and scored for growth. A positively scoring hit is highlighted (box in bottom panel) as an example of a non-looping mutant. (C) Validated hits were analyzed using the cytoscape plugin, BinGO, which created a tree of significantly enriched GO (gene ontology) processes over background. (D) All indicated deletion mutants within the Rpd3L, Rpd3S and Hda1 complexes were re-constructed and spotted onto the indicated media in 10-fold serial dilutions following an overnight culture in YPD to confirm the looping defects identified in the high-throughput screen. Plates were incubated 2–3 days before being imaged. +FOA plates were subsequently replica plated onto SD-URA media to ensure that construct was not lost or mutated. (E) For construct 4, the UAS* was placed in front of the URA3 gene and subsequently integrated at telomere 7L (top). Cell spottings were performed exactly as described in (D) with the indicated genotypes.