期刊
BLOOD
卷 133, 期 23, 页码 2507-2517出版社
AMER SOC HEMATOLOGY
DOI: 10.1182/blood.2018886077
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资金
- FACS facility at Cancer Science Institute of Singapore
- Melamed family
- Leukemia Lymphoma Society of America
- Singapore Ministry of Health's National Medical Research Council (NMRC) under its Singapore Translational Research (STaR) Investigator Award [NMRC/STaR/0021/2014]
- Singapore Ministry of Education Academic Research Fund Tier 2 [MOE2013-T2-2-150]
- NMRC Centre Grant [NMRC/CG/012/2013]
- National Research Foundation Singapore
- Singapore Ministry of Education under its Research Centres of Excellence initiatives
- RNA Biology Centre at the Cancer Science Institute of Singapore
- NUS, as part of funding under the Singapore Ministry of Education's Tier 3 grants [MOE2014-T3-1-006]
CCAAT/enhancer binding protein << (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 16-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 Kru ppel-associated box (KRAB) domain and observed a significant downregulation of transcript and protein levels of CEBPE in cells expressing guide RNA targeting the 16-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 the16-kb enhancer resulted in reduced levels of CEBPE and its target genes and caused a severe block in granulocytic differentiation. Wealso identified binding of CEBPAand CEBPE to the16-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.
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