Please use this identifier to cite or link to this item: http://hdl.handle.net/1946/12422
The aim of this project was to study the roles of Rpt1, Rpt4 and Rpt6 in transcriptional regulation of the GAL network. Therefore, three temperature sentitive degron (td) mutants were created by integrating recombinant plasmids into Saccharomyces cerevisiae chromosomes.
Under galactose induction, degradation of Rpt4 caused a decrease in GAL1 and GAL10 mRNA levels, Rpt1 degradation did not cause any detectable effect and Rpt6 degradation caused an increase in GAL1 and GAL10 mRNA transcribed.
Furthermore, the binding of Tfb3 to the GAL1/10 promoter was studied using formaldehyde-based in vivo cross-linking and chromatin immunoprecipitation (ChIP) in conjunction with mutational and transcriptional analyses. The protein was tagged with a Tandem Affinity Purification (TAP) tag which allows its isolation using an immunoglobulin matrix for studying protein-DNA interactions. In this way, we found that Tfb3 is bound to the GAL1/10 genetic region at position 278326 to 278406 which corresponds to the start of GAL10 gene and its core promoter. We also found that this binding is negatively affected by degradation of Rpt4.