In Vitro Studies of Nucleosome Positioning and Stability at the PHO5 and PHO8 Promoters in Saccharomyces cerevisiae

Acknowledgements First, I want to thank Professor Wolfram Hörz for giving me the opportunity to work his laboratory. During the first years of my work he was a constant source of inspiration. I will remember him not only as a scientific mentor but as a warm and affectionate person who truly cared for his students. I want to thank Philipp Korber for his constant advice and also for his support during especially stressful periods in the lab. I also thank Peter Becker for taking over the role of a formal supervisor of my project and for his big contribution to the establishment and maintenance of a stimulating scientific environment. Dirk Eick I want thank for agreeing on being the second member of my PhD. committee. Thanks to all the people at the Adolf-Butenandt-Institute who made everyday work much more fun and to Hans for his constant support. The work presented in this thesis has been published in the following journal: PHO5 and PHO8 promoters correlates with differential cofactor requirements for chromatin opening. A reprint of this paper can be found in the back of this thesis. The PHO5 and PHO8 genes in yeast provide typical examples for the role of chromatin in promoter regulation. Both genes are regulated by the same transcriptional activator, Pho4, which initiates nucleosome remodeling and transcriptional activation. In spite of this co-regulation, there are important differences in gene activity and in the way promoter chromatin undergoes chromatin remodeling. First, PHO5 belongs to one of the most strongly induced genes in yeast being 10-fold more active than the PHO8 gene (Oshima, 1997; Barbaric et al., 1992). Second, chromatin remodeling at the PHO5 promoter affects four nucleosomes (Almer et al., 1986), whereas only two nucleosomes are afffected at the PHO8 promoter (Barbaric et al., 1992). Third, neither the histone acetyl transferase Gcn5 nor chromatin remodeling complex Swi/Snf seem to be critically required for chromatin remodeling at the PHO5 the complete loss of chromatin remodeling under inducing conditions. Furthermore, Gcn5 is required for full remodeling and transcriptional activation at this promoter (Gregory et al., 1999). Ever since these differences were recognized there have been speculations about the underlying reasons. This work shows that these discrepancies are not a direct consequence of the position or strength of the UASp elements driving the activation of transcription. Instead, these differences result from different stabilities of the two promoter chromatin structures. The basis …