Helix 12 in the human estrogen receptor (hER) is essential for the hER function by overcoming nucleosome repression in yeast

When exogenous human estrogen receptor (hER) binds with estrogen, it can activate transcription of target genes in yeast cells. The estrogen dose-response expression patterns in yeast are very similar to those in human cells. This implies that hER may function in yeast cells via mechanisms similar to those in human cells. In this study, Saccharomyces cerevisiae was used to dissect mechanisms of hER-activated transcription in yeast. The hER contains two transcription activation domains: ER-AF-1 and ER-AF-2 (LBD or HBD). In both human and wild-type yeast cells, hER must bind with estrogen in order to activate transcription. In those cells, ER-AF-2 is independently active upon hormone binding, but ER-AF-1 by itself is inactive. In a mutagenesis screen, we found a mutant strain in which the ER-AF-1 was independently active. It was determined that this mutant strain carried a Tup1 mutation. More interestingly, a small hER fragment ER-AF-0, containing neither ER-AF-1 nor ER-AF-2, Was also fully active in the DeltaTup1 cells. This suggests that in this strain, hormone binding is not required for transcription activation by hER. It is known that the Tup1/Ssn6 complex plays an important role in general transcription repression by protecting histone acetylation sites thus stabilizing nucleosomes. In the DeltaTup1 cells, nucleosomes are known to be unstable because histones can be easily accessed by acetylase and cause nucleosome disassociation. Two point Mutations in helix 12 (H12) in ER-AF-2, which abolished hER function in human cells, also completely abolished hER function in the wild-type yeast cells. This suggested that HI 2 is essential for hER transcription activation function. However, hER with the HI 2 mutation is able to activate transcription in DeltaTup1 cells. This indicates that the normal function of H12 is required for transcription activation by hER only if nucleosomes are not acetylated and are therefore stable. The results of this work suggest that there is a close relationship between hER function and nucleosome remodeling. It also provides insight about H12 activity and its functional relationship with other domains in hER. We propose here that H12 is essential for hER function by recruiting strong nucleosome remodeling proteins to the promoter region thus overcoming nucleosome repression. (C) 2002 Wiley-Liss, Inc.