Naturally-occurring polymorphisms can affect gene expression and thereby underlie human diseases and other important phenotypic traits. A deeper understanding of cis-regulatory variafion will also facilitate the design of better algorithms for predicting cis-regulatory sites and help us elucidate the impact of changes in gene regulafion on morphological evolution. Previously, we analyzed polymorphisms in human and Drosophila microRNA sites and yeast transcription factor binding sites and their relafion to changes in gene expression between individuals in a species, Here we propose to extend this work to cis-regulatory sites that mediate post-transcripfional control via binding of RNA-binding proteins in S. cerevisiae. We use S. cerevisiae as a model system because it offers experimental tractability, arguably the best studied Eukaryofic gene regulatory network and mulfiple sequenced strains. RNA processing is a fundamental cellular process and factors involved in these pathways are conserved in higher Eukaryotes.
Our specific aims for the ROO phase are (1) We will collect the best possible set of motifs for yeast RNA-binding proteins from the literature and by analyzing recent RNA immunoprecipitation (RIP-chip) data and microarray data following knock-out of various RNA-binding proteins. We will test exisfing motif finding tools and try to extend them using regression and conservation techniques, (2) We will predict cis-regulatory sites for RNA binding proteins by combining comparative and populafion genomics data in yeast. We will experimentally validate a subset of the predictions using standard site-directed mutagenesis and qPCR techniques. (3) We will use the recentiy available Digital Gene Expression ? Tag Profiling kit from lllumina to annotate 3'end isoforms In the BY and RM strains and two environmental conditions to help us predict the cis-regulatory sites in Aim 2. If there is at least one gene that is differenfially polyadenylated between BY and RM, we will map cis- and trans- regulators of alternative polyadenylation using either the well-studied BY/RM segregant lines or a panel of recently sequenced wild isolates. Our long-term goal is to use computafional and experimental approaches to link sequence polymorphisms and gene expression changes to morphological change.
Many human diseases are partly caused by the aberrant regulation of genes. Identifying the genefic mutations responsible forthe changes in control ofthese genes is the first step towards diagnosing and curing these diseases. The aim of this project is to develop and validate computafional methods that can eventually be used to create a comprehensive catalogue of gene regulatory variation in the human genome.
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