Activation of transcription is the ultimate endpoint for many signal transduction and developmental pathways, and understanding the mechanism of activation is a key to understanding gene regulation. From previous studies, it is clear that disruption of normal gene regulation by mutations in gene- specific transcription activators and coactivators can lead to cancer and other diseases. The broad long- term objectives of this proposal are to determine the mechanisms used by gene-specific activators and coactivators to regulate RNA polymerase II transcription. The proposed work will provide a basis for understanding gene regulation in normal and diseased states at the molecular level.
The specific aims of this work utilize biochemical, structural, and molecular genetic methods to examine the direct targets of two activation domains and two coactivators. We will examine the structure of several acidic activator-coactivator complexes to understand how activators specifically recognize their targets, common principals of activator-target recognition and, more generally, the function of intrinsically disordered proteins. We will examine the interaction of the SAGA coactivator with TBP (TATA binding protein) and how this interaction is regulated by acetylation. Using a unique set of crosslinking reagents, we will examine the direct targets of the Mediator coactivator complex within the transcription machinery. In all cases, we will use yeast molecular genetics to test the functional significance of our biochemical results. Combined, our results will lead to a molecular model for how activators recognize their coactivator targets and how coactivators stimulate gene expression by direct interaction with the transcription machinery.

Public Health Relevance

Project Narrative The objective of this research is to understand the mechanism and regulation of transcription, the process of mRNA synthesis. Regulation of transcription is one of the key steps in control of cell growth, differentiation, and development, and defects in transcription directly contribute to many human illnesses. Understanding the mechanism of transcription and its regulation will form the basis for understanding the molecular defects in transcription disorders leading to many types of cancer, as well as heart disease, neurological disorders, and birth defects.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM075114-08
Application #
8413436
Study Section
Molecular Genetics A Study Section (MGA)
Program Officer
Sledjeski, Darren D
Project Start
2005-09-01
Project End
2015-01-31
Budget Start
2013-02-01
Budget End
2015-01-31
Support Year
8
Fiscal Year
2013
Total Cost
$584,635
Indirect Cost
$251,399
Name
Fred Hutchinson Cancer Research Center
Department
Type
DUNS #
078200995
City
Seattle
State
WA
Country
United States
Zip Code
98109
Hahn, Steven (2014) Ellis Englesberg and the discovery of positive control in gene regulation. Genetics 198:455-60
Warfield, Linda; Tuttle, Lisa M; Pacheco, Derek et al. (2014) A sequence-specific transcription activator motif and powerful synthetic variants that bind Mediator using a fuzzy protein interface. Proc Natl Acad Sci U S A 111:E3506-13
Gr├╝nberg, Sebastian; Hahn, Steven (2013) Structural insights into transcription initiation by RNA polymerase II. Trends Biochem Sci 38:603-11
Knutson, Bruce A; Hahn, Steven (2011) Domains of Tra1 important for activator recruitment and transcription coactivator functions of SAGA and NuA4 complexes. Mol Cell Biol 31:818-31
Hahn, Steven; Young, Elton T (2011) Transcriptional regulation in Saccharomyces cerevisiae: transcription factor regulation and function, mechanisms of initiation, and roles of activators and coactivators. Genetics 189:705-36
Herbig, Eric; Warfield, Linda; Fish, Lisa et al. (2010) Mechanism of Mediator recruitment by tandem Gcn4 activation domains and three Gal11 activator-binding domains. Mol Cell Biol 30:2376-90
Knutson, Bruce A (2010) Insights into the domain and repeat architecture of target of rapamycin. J Struct Biol 170:354-63
Mohibullah, Neeman; Hahn, Steven (2008) Site-specific cross-linking of TBP in vivo and in vitro reveals a direct functional interaction with the SAGA subunit Spt3. Genes Dev 22:2994-3006
Hahn, Steven (2008) Transcriptional regulation. Meeting on regulatory mechanisms in eukaryotic transcription. EMBO Rep 9:612-6