Molecular Genetic Analysis of Yeast TAFs

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DESCRIPTION (provided by applicant): The goal of our research is to elucidate the molecular mechanisms controlling eukaryotic mRNA production by the enzyme RNA Polymerase II. mRNA serves as the blueprint, or template, for protein synthesis. Defects in mRNA and protein biosynthesis lead to serious human disease such as cancer, cardiovascular disease, diabetes, birth defects and immunodeficiency. Thus, a comprehensive knowledge of the processes controlling mRNA synthesis is essential if we are to truly understand both normal and pathological states, and ultimately intervene to cure disease. Our work utilizes the genetically tractable baker's yeast as a model for elucidating the detailed roles that the highly evolutionarily conserved transcription factor TFIID plays in the complex process of mRNA synthesis, or transcription, by RNA Polymerase II. TFIID is made up of 15 distinct protein subunits and the holo-TFIID complex controls the transcription of over 90% of all mRNA encoding genes in both yeasts and humans. Particular emphasis will be placed on studying how two transcription factors, Repressor Activator Protein 1 (Rap1p) and SBF, collaborate with TFIID to modulate activation of the genes encoding ribosomal protein-encoding genes (Rap1p) and cell-cycle S-phase-dependent mRNA gene transcription (SBF). We will use a multifaceted approach that combines biochemical, biophysical, cell biological and genetic methods to examine the interplay of these transcription factors with each other, gene DNA, the gene-specific DNA-binding regulatory factors Rap1p and SBF, and the RNA polymerase II enzyme that actually synthesizes mRNA, and finally, as appropriate chromatin-modifying coregulators. Successful completion of these experiments will increase our understanding mRNA gene transcription mechanisms. Given that all of these regulatory processes are evolutionarily conserved, our work in the yeast model system will provide insights into human mRNA gene transcription control mechanisms, and hence ultimately human disease.