Published online 4 June 2010 Nucleic Acids Research, 2010, Vol. 38, No. 19 6363–6374 doi:10.1093/nar/gkq490 Transcriptional regulation of Elf-1: locus-wide analysis reveals four distinct promoters, a tissue-specific enhancer, control by PU.1 and the importance of Elf-1 downregulation for erythroid maturation Fernando J. Calero-Nieto*, Andrew D. Wood, Nicola K. Wilson, Sarah Kinston, Josette-Rene´ e Landry and Berthold G o¨ttgens* Department of Haematology, Cambridge Institute for Medical Research, Cambridge University, Hills Road, Cambridge, CB2 0XY, UK Received March 9, 2010; Revised May 11, 2010; Accepted May 14, 2010 ABSTRACT Ets transcription factors play important roles during the development and maintenance of the haemato- poietic system. One such factor, Elf-1 (E74-like factor 1) controls the expression of multiple essen- tial haematopoietic regulators including Scl/Tal1, Lmo2 and PU.1. However, to integrate Elf-1 into the wider regulatory hierarchies controlling haem- atopoietic development and differentiation, regula- tory elements as well as upstream regulators of Elf-1 need to be identified. Here, we have used locus-wide comparative genomic analysis coupled with chromatin immunoprecipitation (ChIP-chip) assays which resulted in the identification of five distinct regulatory regions directing expression of Elf-1. Further, ChIP-chip assays followed by func- tional validation demonstrated that the key haem- atopoietic transcription factor PU.1 is a major upstream regulator of Elf-1. Finally, overexpression studies in a well-characterized erythroid differenti- ation assay from primary murine fetal liver cells demonstrated that Elf-1 downregulation is neces- sary for terminal erythroid differentiation. Given the known activation of PU.1 by Elf-1 and our newly identified reciprocal activation of Elf-1 by PU.1, identification of an inhibitory role for Elf-1 has sig- nificant implications for our understanding of how PU.1 controls myeloid–erythroid differentiation. Our findings therefore not only represent the first report of Elf-1 regulation but also enhance our understand- ing of the wider regulatory networks that control haematopoiesis. INTRODUCTION The formation and subsequent diﬀerentiation of haemato- poietic stem cells (HSCs) is tightly controlled requiring the sequential and coordinated action of multiple transcrip- tion factors. The Ets family is one of the largest families of transcriptional regulators involved in the regulation of haematopoiesis. All Ets proteins contain a region of conserved sequence, corresponding to the ETS domain, which constitutes the DNA binding domain and recog- nizes a core GGAW sequence in the major groove of DNA. Binding of ETS proteins to this sequence is often facilitated by synergistic interaction with transcriptional partners on composite DNA elements (1). The Ets-transcription factor Elf-1 was originally cloned from a human T-cell cDNA library by hybridization with a probe encoding the DNA binding domain (ETS domain) of the human Ets-1 cDNA (2). Based on its preferential expression in embryonic lymphoid organs (thymus and spleen), a wide variety of epithelial cells and fetal liver (3) as well as in adult haematopoietic tissues, including thymus, spleen and bone marrow (4,5), Elf-1 emerged as a potential key regulator of haematopoietic gene *To whom correspondence should be addressed. Tel: +44 1223 336822; Fax: +44 1223 762670; Email: fjc28@cam.ac.uk Correspondence may also be addressed to Berthold Go¨ ttgens. Tel: +44 1223 336829; Fax: +44 1223 762670; Email: bg200@cam.ac.uk Present address: Josette-Rene´ e Landry, Institut de recherche en immunologie et en cance´ rologie, Universite´ de Montre´ al, C.P. 6128, succursale Centre-ville, Montre´ al, Que´ bec, H3C 3J7, Canada. The Author(s) 2010. Published by Oxford University Press. This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/ by-nc/2.5), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.