Genome-Wide Identification of Cis-Regulatory Sequences Controlling Blood and Endothelial Development.

Haematopoiesis has long served as a paradigm for adult stem cell systems and studies over the last 20 years have established that transcriptional control is central to the specification and subsequent differentiation of haematopoietic stem cells (HSCs). With many of the key transcription factors known, haematopoiesis provides a powerful cellular system for the analysis of mammalian gene regulatory networks. The key missing ingredient, particularly for the stem and progenitor cell stages, is a set of experimentally validated gene regulatory regions together with a molecular understanding of their biological activity. Despite progress in lower organisms, genome-wide computational identification of mammalian cis-regulatory sequences has been hindered by increased genomic complexity and cumbersome transgenic assays. In order to identify transcriptional control regions active during early embryonic haematopoietic development, we have developed a strategy that is based upon a haematopoietic stem cell enhancer (+19 enhancer) identified downstream of the SCL transcription factor, a key regulator of haematopoietic stem cell formation. Starting with this well-characterised blood stem cell enhancer, we have developed computational tools which allow genome-wide identification of functionally related enhancers. This approach has been used to identify novel enhancers involved in the regulation of early blood and endothelial development, and which exhibit predicted biological activity in vitro and in transgenic mice. Transcription factors binding to these enhancers have been identified by chromatin immunoprecipitation. Our data allow the construction of an experimentally verified nascent transcriptional network, which controls the development of blood and endothelium, and which so far contains the SCL, GATA-2, Fli-1, Elf-1 and Hex transcription factors. Additional candidate members of the network have been identified and are being subjected to functional validation. This approach represents a widely applicable strategy for characterising the transcriptional networks controlling a broad range of mammalian developmental programmes.