Nucleosome Stability at the Yeast PHO5 and PHO8 Promoters Correlates with Differential Cofactor Requirements for Chromatin Opening

Abstract

The coregulated PHO5 and PHO8 genes in Saccharomyces cerevisiae provide typical examples for the role of chromatin in promoter regulation. It has been a long-standing question why the cofactors Snf2 and Gcn5 are essential for full induction of PHO8 but dispensable for opening of the PHO5 promoter. We show that this discrepancy may result from different stabilities of the two promoter chromatin structures. To test this hypothesis, we used our recently established yeast extract in vitro chromatin assembly system, which generates the characteristic PHO5 promoter chromatin. Here we show that this system also assembles the native PHO8 promoter nucleosome pattern. Remarkably, the positioning information for both native patterns is specific to the yeast extract. Salt gradient dialysis or Drosophila embryo extract does not support proper nucleosome positioning unless supplemented with yeast extract. By competitive assemblies in the yeast extract system we show that the PHO8 promoter has greater nucleosome positioning power and that the properly positioned nucleosomes are more stable than those at the PHO5 promoter. Thus we provide evidence for the correlation of inherently more stable chromatin with stricter cofactor requirements.

ACKNOWLEDGMENTS

We are grateful to Andrea Schmid and Dorothea Blaschke for expert technical assistance, to Peter Becker for critically reading the manuscript, to Gregor Gilfillan for advice on English style, and to all members of the Hörz group for helpful discussions.

This work was supported by the Deutsche Forschungsgemeinschaft (Transregio 5) and the 6th Framework Programme of the European Union (Epigenome Network of Excellence). C.B.H. was supported by Familien Hede Nielsens Fond and Harboefonden.