Temporal and tissue-specific gene expression in mammals depends on complex interactions between transcriptional regulatory proteins and cis-elements such as promoter, enhancers and insulators. Previous large-scale efforts have produced an excellent catalog of transcriptional start sites for most mammalian genes but the mechanisms that control the activation of each promoter in specific cell types remains largely unknown. To better understand the regulatory mechanisms of tissue- and cell type-specific gene expression, it is important to characterize the activities of each promoter in specific cell types, and identify the cis-regulatory elements including enhancers and insulators for each gene. Here, we propose to use the laboratory mouse as a model system and conduct genome-wide analysis of active promoters, enhancers and insulator elements in a panel of embryonic and adult tissues with medical relevance. Specifically, we will (1) identify the active promoters in the mouse genome in a representative set of embryonic and adult tissues;(2) identify potential enhancers and insulator elements in the mouse genome in the same embryonic and adult tissues;(3) identify and characterize tissue-specific promoters and enhancers in the mouse genome;and (4) validate the function of a select set of identified promoters and enhancers. The proposed study, if completed, will result in a comprehensive map of promoters, potential enhancers and insulators throughout the mouse genome. The resource will provide a foundation for analyzing the gene regulatory networks in the mouse cells, and guide the functional annotation of the mouse genome. The results will also help understand the evolution of cis-regulatory sequences, when compared to similar results to be made available as the human ENCODE project progresses. The laboratory mouse represents an important model system for understanding human biology and the molecular basis of human diseases. As a reference to the human genome, the mouse genome sequence has proved extremely valuable in gene annotation. The utility of the laboratory mouse as a model system is currently limited by the lack of understanding of gene regulatory mechanisms that control both common and species-specific gene expression programs in mouse cells. In this project, we propose to define the set of genomic sequences known as cis-regulatory elements in the mouse genome. These regulatory DNA consists of promoters, enhancers, insulators and other regulatory sequences. As a key step towards understanding the gene regulatory mechanisms in mammalian cells, we will produce a comprehensive map of promoters, enhancers and insulators in the mouse genome. We will use a newly developed, high throughput experimental strategy to identify these sequences that are engaged in gene activation in the mouse embryonic stem cells, embryonic fibroblasts, and a panel of embryonic and adult tissues. We will identify tissue specific promoters and enhancers, and characterize the regulatory mechanisms that control the gene expression programs in the specific tissues. Completion of the proposed research is expected to improve our knowledge of the gene regulatory mechanisms in mammalian cells, and provide a reference for understanding the same process in human beings.
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