Large-scale efforts are underway to systematically map transcription factors binding sites throughout the human genome. The ENCODE project has focused its initial attention on two cell lines, 1) K562 cells, a myeloid precursor cell line and 2) GM12878, a lymphoblastoid cell line, and our laboratory has mapped the binding sites of a large number of transcription factor expressed in these cells. To study their conservation and help provide functional information into these binding sites and to determine if these sites are occupied in vivo, we propose two types of studies. First, we will map the binding sites of at least 30 transcription factor orthologs that have been analyzed in the human ENCODE project in mouse MEL and CH12 cells which are analogous to K562 and GM12878 cells, respectively. Second, we will map the binding sites of Pol II and nine other factors in cells differentiated from human CD34+ cells and primary erythroid mouse cells. These studies will determine which transcription factor binding sites and gene targets are conserved in vertebrates and which are species-specific as well as determine the extent to which targets mapped in cultured cell lines reflect in vivo binding sites. The information from these studies will be deposited into public databases and is expected to be extremely valuable to the large mouse and human genetic communities. PUBLIC HEALTH RELEVANCE: The ENCODE project has produced relatively large amounts of data on transcription factor binding and RNA expression in a limited number of human cell lines. We propose to extend these results by obtaining mouse cell lines at similar states of differentiation to human cell lines. We will then duplicate the experiments that have been done in human cells, and locate control elements based on sequence conservation and similarities in factor binding between the two species. We will also determine if elements identified in vitro are occupied in cells isolated from organisms.