Eukaryotes dedicate hundreds (and in some cases thousands) of proteins towards the regulation of gene expression. Yet very little is known about how all of these proteins coordinate their behavior at the many thousands of genes that comprise a typical genome. Little is known about how this coordination changes as cells reprogram their genome in response to signaling events including environmental stress. The work proposed here uses Saccharomyces cerevisiae as a model cellular system to undertake a broad survey of where transcriptional regulatory proteins are located throughout the genome, and where they move to when the genome is reprogrammed by environmental signals, such as heat shock and other stresses. Heat shock provides a rapid and simple programming event for the cell. Preliminary studies on this project have already revealed novel insights into gene regulation by demonstrating that many genes undergo partial assembly of the transcription machinery at promoters. Partial complexes await signaling events that drive them into full assembly. The location of a wide range of proteins involved in transcription will be evaluated by chromatin immunoprecipitation assays in which microarrays are used to detected genome-wide binding events (so called chlP-chip). Location will be assessed under normal growth conditions and under a wide range of environmental stresses, with particular emphasis on heat shock. Relationships among binding events will provide new insights into transcription complex assembly and regulation. Additional mechanistic insight will be provided through genome-wide biochemical dissection of native transcription complexes isolated from cells. Our cells are constantly faced with environmental extremes, involving temperature, starvation, radiation and harmful chemicals. How we deal with this stress depends upon the action of our transcription machinery. Therefore, a broad understanding of how our transcription machinery works in the face of various stresses is essential for a physiological understanding of human health.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Research Project (R01)
Project #
5R01ES013768-04
Application #
7754405
Study Section
Molecular Genetics B Study Section (MGB)
Program Officer
Balshaw, David M
Project Start
2007-01-01
Project End
2010-12-31
Budget Start
2010-01-01
Budget End
2010-12-31
Support Year
4
Fiscal Year
2010
Total Cost
$300,446
Indirect Cost
Name
Pennsylvania State University
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
003403953
City
University Park
State
PA
Country
United States
Zip Code
16802
Mahony, Shaun; Pugh, B Franklin (2015) Protein-DNA binding in high-resolution. Crit Rev Biochem Mol Biol 50:269-83
Chang, Gue Su; Chen, Xiangyun Amy; Park, Bongsoo et al. (2014) A comprehensive and high-resolution genome-wide response of p53 to stress. Cell Rep 8:514-27
Nakahashi, Hirotaka; Kieffer Kwon, Kyong-Rim; Resch, Wolfgang et al. (2013) A genome-wide map of CTCF multivalency redefines the CTCF code. Cell Rep 3:1678-1689
Li, Jian; Liu, Yingyun; Rhee, Ho Sung et al. (2013) Kinetic competition between elongation rate and binding of NELF controls promoter-proximal pausing. Mol Cell 50:711-22
Rhee, Ho Sung; Pugh, B Franklin (2012) ChIP-exo method for identifying genomic location of DNA-binding proteins with near-single-nucleotide accuracy. Curr Protoc Mol Biol Chapter 21:Unit 21.24
Ghosh, Sujana; Pugh, B Franklin (2011) Sequential recruitment of SAGA and TFIID in a genomic response to DNA damage in Saccharomyces cerevisiae. Mol Cell Biol 31:190-202
Rhee, Ho Sung; Pugh, B Franklin (2011) Comprehensive genome-wide protein-DNA interactions detected at single-nucleotide resolution. Cell 147:1408-19
Venters, Bryan J; Wachi, Shinichiro; Mavrich, Travis N et al. (2011) A comprehensive genomic binding map of gene and chromatin regulatory proteins in Saccharomyces. Mol Cell 41:480-92
Samorodnitsky, Eric; Pugh, B Franklin (2010) Genome-wide modeling of transcription preinitiation complex disassembly mechanisms using ChIP-chip data. PLoS Comput Biol 6:e1000733
Venters, Bryan J; Pugh, B Franklin (2009) A canonical promoter organization of the transcription machinery and its regulators in the Saccharomyces genome. Genome Res 19:360-71

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