Chromatin dynamics at the Saccharomyces cerevisiae PHO5 promoter

In eukaryotes, the organization of DNA into chromatin is a primary determinant of gene expression. Positioned nucleosomes in promoter regions are frequently found to regulate gene expression by obstructing the accessibility of cis-regulatory elements in DNA to trans-factors. This dissertation focuses on the chromatin structure and remodeling program at the S. cerevisiae PHO5 promoter, extending the use of DNA methyltransferases as in vivo probes of chromatin structure. Our studies address the diversity of histone-DNA interactions in vivo by examining nucleosome conformational stabilities at the PHO5 promoter. We present high-resolution chromatin structural mapping of the promoter, required to relate in vivo site accessibility to nucleosome stability and show that the PHO5 promoter nucleosomes have different accessibilities. We show a correlation between DNA curvature and nucleosome positioning, which is consistent with the observed differences in accessibility/stability. Kinetic analyses of the chromatin remodeling program at PHO5 show that nucleosomes proximal to the enhancer are disrupted preferentially and prior to those more distal, demonstrating bidirectional and finite propagation of chromatin remodeling from bound activators and providing a novel mechanism by which transactivation at a distance occurs.