? Mature neutrophils arise from the hematopoietic stem cell via a series of commitment steps. The appearance of the secondary granule proteins (SGP) lactoferrin (LF), transcobalamin I (TCI), neutrophil collagenase (NC) and neutrophil gelatinase (NG) marks the commitment to terminal neutrophil differentiation. C/EBPepsilon (C/EBPe) plays a critical role in the coordinate upregulation of SGP genes. Disruption of the C/EBPe gene in mice leads to morphologic and functional defects in neutrophil maturation with a defective transition from the promyelocyte to the myelocyte stage. The neutrophils have bilobed nuclei, abnormal respiratory burst activity, and impaired chemotaxis and bactericidal activity. They lack specific granules and fail to express mRNAs encoding for secondary and tertiary granule content proteins. The mice die within 3-5 months of infection or from complications of """"""""myeloproliferation"""""""". Phenotypic and functional defects of the C/EBPe -/- mice closely parallel those in patients with secondary granule deficiency. We have made 2 different cell lines from the bone marrow of the C/EBPe -/- mice and corresponding wildtype littermates that mimic the morphologic and functional defects in the knockout mice. Using these cell lines and primary marrow cells, we propose to further characterize the transcriptional regulation of terminal neutrophil differentiation and specifically the role of C/EBPe in the neutrophil maturation program.
Our specific aims are: 1) To complete the characterization of the newly generated cell lines and establish them as a faithful model of the C/EBPE -/- phenotye: 2) To identify downstream targets of C/EBPepsilon through microarray analysis of the cell lines and primary C/EBPe +/+ and -/- bone marrow; and 3) To rescue the C/EBPepsilon -/- phenotype by retroviral transduction with candidate genes identified in specif aim 2 . Genes will be transferred first into the C/EBPe -/- cell line to determine reversal of phenotype. Verified important targets will be transduced into C/EBPe -/- marrow progenitors and transplanted back into C/EBPe -/- mice to assess reversal of phenotype ? ?
| Taylor, Ashley; Tang, Wenwen; Bruscia, Emanuela M et al. (2014) SRF is required for neutrophil migration in response to inflammation. Blood 123:3027-36 |
| Halene, Stephanie; Gao, Yuan; Hahn, Katherine et al. (2010) Serum response factor is an essential transcription factor in megakaryocytic maturation. Blood 116:1942-50 |
| Halene, Stephanie; Gaines, Peter; Sun, Hong et al. (2010) C/EBPepsilon directs granulocytic-vs-monocytic lineage determination and confers chemotactic function via Hlx. Exp Hematol 38:90-103 |
