The broad, long-term objectives of this proposal are to determine at a molecular level the mechanism of transcription initiation by RNA polymerase III and how this relates to the initiation mechanisms for the other nuclear polymerases. This will lead to a better understanding of the transcription initiation machinery and how it is regulated. This is important because the transcription machinery is the ultimate target of many signal transduction pathways and gene-specific regulatory factors. Understanding the fundamental process of gene control will provide a basis for understanding many types of diseases such as cancer, which often result from aberrant regulation of transcription initiation. In addition, other diseases resulting from defects in repair of DNA damage are caused by mutations in components of the transcription machinery.
The specific aims of this project are designed to study through molecular, genetic and biochemical methods, the function of TFIIIB, a multi-subunit component of the Pol III transcription machinery. Dr. Hahn will complete the cloning of the TFIIIB subunit TFIIIB90 and perform a molecular and genetic analysis of its function. Dr. Hahn will continue the genetic and biochemical analysis of BRF1, a second subunit of TFIIIB. He will also use a genetic approach to understand the role of TATA binding protein, a third subunit of TFIIIB, in Pol III transcription Finally, Dr. Hahn has devised a strategy to examine in detail the role of the individual TFIIIB subunits in the process of polymerase recruitment and open complex formation. His work will involve molecular techniques (gene cloning, mutagenesis, and PCR), biochemical fractionation, in vitro transcription, protein biochemistry, and yeast molecular genetics.
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