The Cebpa +37-kb enhancer directs transgene expression to myeloid progenitors and to long-term hematopoietic stem cells

J Leukoc Biol. 2014 Sep;96(3):419-26. doi: 10.1189/jlb.2AB0314-145R. Epub 2014 May 27.


C/EBPα is expressed preferentially in myeloid compared with lymphoid or erythroid cells and directs myeloid lineage specification. C/EBPα is also expressed at lower levels in HSCs and in several nonhematopoietic tissues. The Cebpa gene has a conserved, 450-bp segment at +37 kb that harbors enhancer-specific epigenetic marks and is activate in a myeloid cell line. Herein, we characterize transgenic C57BL/6 mice, in which the Cebpa enhancer and 845-bp promoter regulate a hCD4 reporter. FACS analysis, in vitro colony assays, and in vivo competitive and secondary transplantation revealed that myeloid but not MEPs or lymphoid progenitors and also functional LT-HSCs are found almost exclusively in the Cebpa-hCD4(+) compared with hCD4(-) marrow population. hCD4(+) CMP yielded predominantly myeloid, whereas hCD4(-) CMP generated mainly Meg/E colonies. Providing insight into control of CMP maturation, Cebpa and Pu.1 RNAs were preferentially expressed in hCD4(+) CMP, Scl, Gata2, Gata1, Klf1, Ets1, and Fli1 predominated in hCD4(-) CMP, and Runx1, Myb, HoxA9, and Erg levels were similar in both. Cebpa-hCD4 transgene expression was lacking in multiple nonhematopoietic tissues. In summary, the +37-kb Cebpa enhancer and promoter are sufficient for marrow myeloid progenitor and LT-HSC-specific expression.

Keywords: differentiation; hematopoiesis; myelopoiesis.

Publication types

  • Comparative Study
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bone Marrow Transplantation
  • CCAAT-Enhancer-Binding Proteins / genetics*
  • CCAAT-Enhancer-Binding Proteins / physiology
  • CD4 Antigens / biosynthesis
  • CD4 Antigens / chemistry
  • CD4 Antigens / genetics
  • Cell Lineage
  • Cell Separation
  • Colony-Forming Units Assay
  • Enhancer Elements, Genetic / genetics*
  • Flow Cytometry
  • Gene Expression Regulation / genetics
  • Genes, Reporter
  • Hematopoietic Stem Cells / cytology*
  • Humans
  • Lymphocytes / cytology
  • Lymphopoiesis / genetics
  • Mice, Inbred C57BL
  • Mice, Transgenic
  • Myeloid Cells / cytology*
  • Myelopoiesis / genetics
  • Peptide Fragments / biosynthesis
  • Peptide Fragments / genetics
  • Promoter Regions, Genetic
  • Radiation Chimera
  • Recombinant Fusion Proteins / biosynthesis
  • Transgenes


  • CCAAT-Enhancer-Binding Proteins
  • CD4 Antigens
  • CEBPA protein, human
  • Peptide Fragments
  • Recombinant Fusion Proteins