title: C/EBPepsilon directs granulocytic-vs-monocytic lineage determination and confers chemotactic function via Hlx.
abstract: OBJECTIVE
  Mutations in the CCAAT enhancer binding protein epsilon (C/EBPepsilon) gene have been identified in the cells of patients with neutrophil specific granule deficiency, a rare congenital disorder marked by recurrent bacterial infections. Their neutrophils, in addition to lacking specific granules required for normal respiratory burst activity, also lack normal phagocytosis and chemotaxis. Although the specific granule deficiency phenotype has been replicated in C/EBPepsilon(-/-) (knockout [KO]) mice, the mechanisms by which C/EBPepsilon mutations act to decrease neutrophil function are not entirely clear.
  
  
  METHODS
  In order to determine the role of C/EBPepsilon in neutrophil differentiation and migration, we generated immortalized progenitor cell lines from C/EBPepsilon KO and wild-type mice and performed expression and flow cytometric analysis and functional studies.
  
  
  RESULTS
  Expression of lineage-specific cell surface antigens on our in vitro differentiated cell lines revealed persistent expression of monocytic markers on KO granulocytes. We verified this in primary murine peripheral blood and bone marrow cells. In addition, KO bone marrow had an increase in immature myeloid precursors at the common myeloid progenitor and granulocyte/monocyte progenitor levels, suggesting a critical role for C/EBPepsilon not only in granulocyte maturation beyond the promyelocyte stage, but also in the monocyte/granulocyte lineage decision. We found that restoration of Hlx (H2.0-like homeo box 1) expression, which was decreased in C/EBPepsilon KO cells, rescued chemotaxis, but not the other defects of C/EBPepsilon KO neutrophils.
  
  
  CONCLUSIONS
  We show two new regulatory functions of C/EBPepsilon in myelopoiesis: in the absence of C/EBPepsilon, there is not only incomplete differentiation of granulocytes, but myelopoiesis is disrupted with the appearance of an intermediate cell type with monocyte and granulocyte features, and the neutrophils have abnormal chemotaxis. Restoration of expression of Hlx provides partial recovery of function; it has no effect on neutrophil maturation, but can completely ameliorate the chemotaxis defect in C/EBPepsilon KO cells.
authors.affiliation: Section of Hematology, Department of Internal Medicine and Yale Cancer Center, Yale University School of Medicine, New Haven, CT 06520-8073, USA. Stephanie.halene@yale.edu
authors.forename: Stephanie
authors.initials: S
authors.lastname: Halene
authors.forename: Peter
authors.initials: P
authors.lastname: Gaines
authors.forename: Hong
authors.initials: H
authors.lastname: Sun
authors.forename: Theresa
authors.initials: T
authors.lastname: Zibello
authors.forename: Sharon
authors.initials: S
authors.lastname: Lin
authors.forename: Arati
authors.initials: A
authors.lastname: Khanna-Gupta
authors.forename: Simon C
authors.initials: SC
authors.lastname: Williams
authors.forename: Archibald
authors.initials: A
authors.lastname: Perkins
authors.forename: Diane
authors.initials: D
authors.lastname: Krause
authors.forename: Nancy
authors.initials: N
authors.lastname: Berliner
mesh_terms: D000818:Animals; D001854:Bone Marrow Cells / Q000166:cytology; D022762:CCAAT-Enhancer-Binding Proteins / Q000235:genetics / Q000502:physiology*; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology*; D002460:Cell Line; D002634:Chemotaxis, Leukocyte / Q000502:physiology*; D015870:Gene Expression; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006098:Granulocytes / Q000737:chemistry / Q000166:cytology*; D006412:Hematopoietic Stem Cells / Q000166:cytology; D018398:Homeodomain Proteins / Q000235:genetics / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D009000:Monocytes / Q000166:cytology*; D038042:Myelopoiesis / Q000502:physiology; D009504:Neutrophils / Q000737:chemistry / Q000166:cytology / Q000502:physiology; D019707:Receptors, Chemokine / Q000032:analysis; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D014161:Transduction, Genetic
chemical_list: D022762:CCAAT-Enhancer-Binding Proteins; C489468:Cebpe protein, mouse; C503133:Gr-1 protein, mouse; C489999:Hlx protein, mouse; D018398:Homeodomain Proteins; D019707:Receptors, Chemokine; D014157:Transcription Factors; D016178:Granulocyte-Macrophage Colony-Stimulating Factor