title: The Cebpa +37-kb enhancer directs transgene expression to myeloid progenitors and to long-term hematopoietic stem cells.
abstract: 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.
authors.affiliation: Division of Pediatric Oncology, Johns Hopkins University, Baltimore, Maryland, USA.
authors.forename: Hong
authors.initials: H
authors.lastname: Guo
authors.affiliation: Division of Pediatric Oncology, Johns Hopkins University, Baltimore, Maryland, USA.
authors.forename: Ou
authors.initials: O
authors.lastname: Ma
authors.affiliation: Division of Pediatric Oncology, Johns Hopkins University, Baltimore, Maryland, USA afriedm2@jhmi.edu.
authors.forename: Alan D
authors.initials: AD
authors.lastname: Friedman
mesh_terms: D000818:Animals; D016026:Bone Marrow Transplantation; D022762:CCAAT-Enhancer-Binding Proteins / Q000235:genetics* / Q000502:physiology; D015704:CD4 Antigens / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D019070:Cell Lineage; D002469:Cell Separation; D003114:Colony-Forming Units Assay; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D005434:Flow Cytometry; D005786:Gene Expression Regulation / Q000235:genetics; D017930:Genes, Reporter; D006412:Hematopoietic Stem Cells / Q000166:cytology*; D006801:Humans; D008214:Lymphocytes / Q000166:cytology; D038041:Lymphopoiesis / Q000235:genetics; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D022423:Myeloid Cells / Q000166:cytology*; D038042:Myelopoiesis / Q000235:genetics; D010446:Peptide Fragments / Q000096:biosynthesis / Q000235:genetics; D011401:Promoter Regions, Genetic; D011828:Radiation Chimera; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D019076:Transgenes
chemical_list: D022762:CCAAT-Enhancer-Binding Proteins; D015704:CD4 Antigens; C515848:CEBPA protein, human; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins
keywords: differentiation; hematopoiesis; myelopoiesis