Activation of transcription is the ultimate endpoint for many signal transaction and developmental pathways, and understanding the mechanism of activation is a key to understanding the action of these pathways. From previous studies, it is clear that disruption of normal gene regulation by mutations in gene-specific transcription activators can lead to cancer and other diseases. The broad long-term objectives of this proposal are to determine the mechanisms used by gene-specific transcription factors to activate transcription by RNA Polymerase II (Pol II). 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 will utilize biochemical, structural, and molecular genetic methods to examine the direct targets of several transcription activators and the mechanism whereby contact with these targets stimulates transcription. Using a newly developed method for mapping protein-protein contacts within large complexes, we will identify direct targets of activation domains in several model activators in the presence or absence of chromatin. Biochemical and molecular genetic studies will be conducted to test the relevance of these interactions in transcription. We will extend these studies to examine the targets of a different activator class acting at TATA-less promoters. After mapping these activator-target interactions by hydroxyl radical cleavage and direct protein-protein interaction assays, we will determine the structure of the activation domains in combination with their relevant targets. Finally, by blocking assembly of the Preinitiation Complex at intermediate stages, we will examine the mechanism by which activator contact with 1 of these targets, the SAGA coactivator complex, stimulates transcription. Our proposed work will illuminate important mechanisms and principles of transcriptional regulation.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM075114-01
Application #
6961840
Study Section
Molecular Genetics A Study Section (MGA)
Program Officer
Tompkins, Laurie
Project Start
2005-09-01
Project End
2009-08-31
Budget Start
2005-09-01
Budget End
2006-08-31
Support Year
1
Fiscal Year
2005
Total Cost
$473,082
Indirect Cost
Name
Fred Hutchinson Cancer Research Center
Department
Type
DUNS #
078200995
City
Seattle
State
WA
Country
United States
Zip Code
98109
Bruzzone, Maria Jessica; Grünberg, Sebastian; Kubik, Slawomir et al. (2018) Distinct patterns of histone acetyltransferase and Mediator deployment at yeast protein-coding genes. Genes Dev 32:1252-1265
Pacheco, Derek; Warfield, Linda; Brajcich, Michelle et al. (2018) Transcription Activation Domains of the Yeast Factors Met4 and Ino2: Tandem Activation Domains with Properties Similar to the Yeast Gcn4 Activator. Mol Cell Biol 38:
Tuttle, Lisa M; Pacheco, Derek; Warfield, Linda et al. (2018) Gcn4-Mediator Specificity Is Mediated by a Large and Dynamic Fuzzy Protein-Protein Complex. Cell Rep 22:3251-3264
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
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
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; 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
Han, Yan; Luo, Jie; Ranish, Jeffrey et al. (2014) Architecture of the Saccharomyces cerevisiae SAGA transcription coactivator complex. EMBO J 33:2534-46
Hahn, Steven (2014) Ellis Englesberg and the discovery of positive control in gene regulation. Genetics 198:455-60

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