RUNX1 regulates the CD34 gene in haematopoietic stem cells by mediating interactions with a distal regulatory element

• Published in: The EMBO journal 2011-10, Vol.30 (19), p.4059-4070
• Main Authors: Levantini, Elena, Lee, Sanghoon, Radomska, Hanna S, Hetherington, Christopher J, Alberich-Jorda, Meritxell, Amabile, Giovanni, Zhang, Pu, Gonzalez, David A, Zhang, Junyan, Basseres, Daniela S, Wilson, Nicola K, Koschmieder, Steffen, Huang, Gang, Zhang, Dong-Er, Ebralidze, Alexander K, Bonifer, Constanze, Okuno, Yutaka, Gottgens, Bertie, Tenen, Daniel G
• Format: Article
• Language: English
• Subjects: Animals; Antigens, CD34 - metabolism; Binding sites; Bone Marrow Transplantation; CD34; Chromatin; Chromatin - metabolism; Core Binding Factor Alpha 2 Subunit - genetics; Core Binding Factor Alpha 2 Subunit - metabolism; EMBO09; EMBO11; Fetal Blood - cytology; Gene expression; Gene Expression Regulation; gene regulation; Genotype; haematopoietic stem cell; Hematopoietic Stem Cells - metabolism; HL-60 Cells; Humans; long-range chromatin looping; Mice; Mice, Transgenic; Models, Biological; Molecular biology; Proteins; Regulatory Sequences, Nucleic Acid - genetics; RUNX proteins; Stem cells
• ISSN: 0261-4189; 1460-2075
• DOI: 10.1038/emboj.2011.285

The transcription factor RUNX1 is essential to establish the haematopoietic gene expression programme; however, the mechanism of how it activates transcription of haematopoietic stem cell (HSC) genes is still elusive. Here, we obtained novel insights into RUNX1 function by studying regulation of the human CD34 gene, which is expressed in HSCs. Using transgenic mice carrying human CD34 PAC constructs, we identified a novel downstream regulatory element (DRE), which is bound by RUNX1 and is necessary for human CD34 expression in long‐term (LT)‐HSCs. Conditional deletion of Runx1 in mice harbouring human CD34 promoter–DRE constructs abrogates human CD34 expression. We demonstrate by chromosome conformation capture assays in LT‐HSCs that the DRE physically interacts with the human CD34 promoter. Targeted mutagenesis of RUNX binding sites leads to perturbation of this interaction and decreased human CD34 expression in LT‐HSCs. Overall, our in vivo data provide novel evidence about the role of RUNX1 in mediating interactions between distal and proximal elements of the HSC gene CD34 . This study reveals molecular details of RUNX1 function in haematopoiesis. Employing various in vivo genetic models and chromatin conformation capture (3C) assays in HSCs, RUNX1 is shown to regulate CD34 expression by looping of a newly characterized distal regulatory element to the core promoter.