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Blood. 2019 Jun 6;133(23):2507-2517. doi: 10.1182/blood.2018886077. Epub 2019 Apr 5.

Identification of a novel enhancer of CEBPE essential for granulocytic differentiation.

Author information

1
Cancer Science Institute of Singapore and.
2
Department of Medicine, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
3
Division of Hematology/Oncology, Cedars-Sinai Medical Center, UCLA School of Medicine, Los Angeles, CA; and.
4
Department of Hematology-Oncology, National University Cancer Institute of Singapore, National University Hospital, Singapore.

Abstract

CCAAT/enhancer binding protein ε (CEBPE) is an essential transcription factor for granulocytic differentiation. Mutations of CEBPE occur in individuals with neutrophil-specific granule deficiency (SGD), which is characterized by defects in neutrophil maturation. Cebpe-knockout mice also exhibit defects in terminal differentiation of granulocytes, a phenotype reminiscent of SGD. Analysis of DNase I hypersensitive sites sequencing data revealed an open chromatin region 6 kb downstream of the transcriptional start site of Cebpe in murine myeloid cells. We identified an interaction between this +6-kb region and the core promoter of Cebpe using circular chromosome conformation capture sequencing (4C-seq). To understand the role of this putative enhancer in transcriptional regulation of Cebpe, we targeted it using catalytically inactive Cas9 fused to Krüppel-associated box (KRAB) domain and observed a significant downregulation of transcript and protein levels of CEBPE in cells expressing guide RNA targeting the +6-kb region. To further investigate the role of this novel enhancer further in myelopoiesis, we generated mice with deletion of this region using CRISPR/Cas9 technology. Germline deletion of the +6-kb enhancer resulted in reduced levels of CEBPE and its target genes and caused a severe block in granulocytic differentiation. We also identified binding of CEBPA and CEBPE to the +6-kb enhancer, which suggests their role in regulating the expression of Cebpe In summary, we have identified a novel enhancer crucial for regulating expression of Cebpe and required for normal granulocytic differentiation.

PMID:
30952671
DOI:
10.1182/blood.2018886077
[Indexed for MEDLINE]

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