The Roles of Flanking DNA and Transcriptional Activators in Regulating SAGA‐Mediated Nucleosome Acetylation

Abstract

The SAGA family of transcriptional coactivators are prototypical nucleosome acetyltransferase complexes that regulate multiple steps of gene transcription. The size and complexity of both the SAGA enzyme and the chromatin substrate provide opportunities to regulate the acetylation process. To better probe this regulation, we have characterized the binding interactions and kinetics of acetylation using different nucleosomal substrates and the full SAGA complex purified from budding yeast. We find that SAGA‐mediated nucleosome acetylation proceeds through a multiple‐turnover burst phase, suggesting rapid nucleosome acetylation at multiple sites prior to nucleosome release. The rate of nucleosome acetylation is affected by DNA flanking the nucleosome, both by facilitating the binding of SAGA and by accelerating acetylation turnover. This stimulation requires flanking DNA on both sides of the nucleosome, where one side needs to be longer than 15 base pairs. Gal4‐VP16 activator can also augment nucleosome acetylation. However, contrary to our expectations, this stimulation does not appear to occur by stabilizing the binding of SAGA toward nucleosomes containing an activator binding site. Instead, stimulation of nucleosome acetylation occurs by increasing acetylation turnover by SAGA. Altogether, these studies uncover several novel mechanisms of SAGA regulation by chromatin substrates.Support or Funding InformationThis work was supported by American Cancer Society Research Scholar Grant 1206501 to M. A. S.‐K. Work by S. J. O. was supported in part by a Gary Roewe Faculty Research Award.