Biochemical, Genomic and Computational Analysis of Transcriptional Repression

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Project Summary Transcriptional repressors represent the necessary counterweight to transcriptional activators in metazoan development. The mechanisms of transcriptional repression have been intensively investigated, but we lack crucial insights into how repressors act at a mechanistic level across many targets in the genome. In particular, the coordinated recruiting of transcriptional co-repressors can generate diverse effects on chromatin structure and modification, and we still lack insights on the functional significance of many changes that can be measured in `omics studies. To develop key insights into eukaryotic transcriptional regulatory mechanisms, we use the natural setting of the Drosophila embryo to identify basic biochemical processes in a developmental setting, where differential gene expression is used to drive the developmental fate of particular cells and tissues. In this proposal, 1) we will use genome-wide methods developed in our laboratory to identify direct biochemical, chromatin-based processes that are directed by a set of five endogenous transcriptional regulators that repress through short-range and long-range mechanisms. 2) We will study the in vivo activity of wild-type and mutant repression complexes to identify the contributions of distinct transcriptional co-repressors Groucho and CtBP, testing their contributions to quantitative and/or qualitative effects in chromatin modifications and gene regulation. 3) To identify the cis-regulatory context in which transcriptional repressors act on different enhancers, we will quantitatively measure and mathematically model the output of specific enhancers to uncover the fundamental quantitative properties of specific classes of repressors interacting with activators – i.e. the common “rules” by which these proteins interact on many target genes. The three interrelated aims will provide predictive tools for interpretation of genomic cis regulatory content of the metazoan genome, providing essential underpinnings for studies of evolution and disease in higher eukaryotes.