Large Scale Identification and Characterization of Hsf1-mediated Heat Shock Respo

Princeton University, Princeton NJ

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Abstract

DESCRIPTION (provided by applicant): The ability to sense and respond to changing environmental conditions is a fundamental aspect of all biological systems. The conserved heat shock response in Saccharomyces cerevisiae has proven to be a paradigm for general eukaryotic stress responses. Despite much analysis at the level of gene expression and the functions of those expressed genes, very little is known about the cellular sensor or potential signal transduction pathway that activates the conserved heat shock transcription factor, the Hsf1 protein (referred to as Hsf1). The research outlined in this proposal seeks to identify and characterize proteins that act as regulators of Hsf1. A preliminary screen was performed and resulted in identification of 11 potential positive Hsf1 regulators. The first part of this proposal is dedicated to examining each of these genes for their functional and conserved role in the Hsf1-mediated heat shock response. This will be accomplished using a combination of classic genetic and biochemical techniques (such as deletion analysis and Western blot analysis) and systems-level biological techniques (such as competitive fitness assays and microarrays). The second part of this proposal is dedicated to developing a versatile, quantitative system for identification of both positive and negative regulators of transcription factors, which will be used to further identify Hsf1 regulators. This will be accomplished using common recombinant DNA technology in addition to fluorescent activated cell sorting (FACS) and barcode DNA sequencing using next generation sequencing technology (Illumina Genome Analyzer II). Overall, this work will significantly increase our knowledge of the heat shock response, and of eukaryotic stress- responsive signal transduction, in general.