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.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM053451-04
Application #
2685079
Study Section
Molecular Biology Study Section (MBY)
Program Officer
Tompkins, Laurie
Project Start
1995-09-30
Project End
2000-03-31
Budget Start
1998-04-01
Budget End
1999-03-31
Support Year
4
Fiscal Year
1998
Total Cost
Indirect Cost
Name
Fred Hutchinson Cancer Research Center
Department
Type
DUNS #
075524595
City
Seattle
State
WA
Country
United States
Zip Code
98109
Donczew, Rafal; Hahn, Steven (2018) Mechanistic Differences in Transcription Initiation at TATA-Less and TATA-Containing Promoters. Mol Cell Biol 38:
Grünberg, Sebastian; Zentner, Gabriel E (2017) Genome-wide Mapping of Protein-DNA Interactions with ChEC-seq in Saccharomyces cerevisiae. J Vis Exp :
Grünberg, Sebastian; Zentner, Gabriel E (2017) Genome-wide characterization of Mediator recruitment, function, and regulation. Transcription 8:169-174
Warfield, Linda; Ramachandran, Srinivas; Baptista, Tiago et al. (2017) Transcription of Nearly All Yeast RNA Polymerase II-Transcribed Genes Is Dependent on Transcription Factor TFIID. Mol Cell 68:118-129.e5
Baptista, Tiago; Grünberg, Sebastian; Minoungou, Nadège et al. (2017) SAGA Is a General Cofactor for RNA Polymerase II Transcription. Mol Cell 68:130-143.e5
Tomko, Eric J; Fishburn, James; Hahn, Steven et al. (2017) TFIIH generates a six-base-pair open complex during RNAP II transcription initiation and start-site scanning. Nat Struct Mol Biol 24:1139-1145
Fishburn, James; Galburt, Eric; Hahn, Steven (2016) Transcription Start Site Scanning and the Requirement for ATP during Transcription Initiation by RNA Polymerase II. J Biol Chem 291:13040-7
Grünberg, Sebastian; Henikoff, Steven; Hahn, Steven et al. (2016) Mediator binding to UASs is broadly uncoupled from transcription and cooperative with TFIID recruitment to promoters. EMBO J 35:2435-2446
Warfield, Linda; Luo, Jie; Ranish, Jeffrey et al. (2016) Function of Conserved Topological Regions within the Saccharomyces cerevisiae Basal Transcription Factor TFIIH. Mol Cell Biol 36:2464-75
Fishburn, James; Tomko, Eric; Galburt, Eric et al. (2015) Double-stranded DNA translocase activity of transcription factor TFIIH and the mechanism of RNA polymerase II open complex formation. Proc Natl Acad Sci U S A 112:3961-6

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