DNA is transformed into a living genome by processes that occur in the cells nucleus, such as transcription, replication, recombination, and DNA repair. These hallmarks of life are controlled by regulatory networks that ultimately modulate, as a function of time and environment, just two aspects of DNA-dependent enzymes: the level of their activity, and where they act. Therefore, a critical part of understanding the mechanism and logic of cellular regulatory networks is a comprehensive understanding of where enzymes and their regulatory proteins interact with the genome in vivo. High-resolution, genome-wide maps of protein-DNA interactions can be made by immunoprecipitating specific protein-DNA complexes and determining the genomic location of the lP-enriched DNA by microarray hybridization. With this information, we can identify the genomic features that specify protein binding, and simultaneously identify genes or other chromosomal elements whose function is affected by the binding. The broad goal of this proposal is to use this and other methods to map the regulatory functions of non-coding sequences onto the entire genome of the yeast Saccharomyces cerevisiae, the most tractable eukaryote, and then to extend this approach to a model metazoan, Caenorhabditis elegans. How do DNA-binding proteins attain their genome-wide target specificity in vivo? This question arises from previous work showing that the transcription factor Rap1 binds to non-coding regions upstream of genes in preference to the coding regions, even though both regions contain strong consensus Rap1 binding sites. The proposed yeast experiments use Rap1 and its associated DNA-binding proteins as a model system to investigate the unaccounted-for determinants of in vivo binding specificity. In addition, by correlating the genome-wide position of Rap1 and its cofactors with expression level of downstream genes, we aim to discover how combinations of proteins assembled at promoters code for specific transcriptional outputs. Currently, I am a third-year Helen Hay Whitney postdoctoral fellow in the laboratory of Patrick Brown at Stanford University. I recently accepted an assistant professor position (tenure-track) at the University of North Carolina in Chapel Hill in the Department of Biology and Carolina Center for the Genome Sciences, starting in June, 2002. Therefore, this application is for only the Faculty Transition Phase of this award. UNC offers an excellent research environment for the proposed work, providing 1000 sq. ft. of wet bench lab space, a state-of-the-ad DNA microarrayer, and generous start-up funding. The close alignment between my research interests and those of colleagues at UNC ensures a rich environment for collaboration and the exchange of knowledge. The award would greatly facilitate the establishment of a productive laboratory engaged in cutting-edge genomics research.
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