Impaired granulopoiesis, myelodysplasia, and early lethality in CCAAT/enhancer binding protein -deficient mice (original) (raw)
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Journal of Biological Chemistry, 2006
CCAAT/enhancer-binding protein ⑀ (C/EBP⑀) plays a critical role in terminal myeloid differentiation. Differentiation is an integrated process of cell cycle arrest, morphological change, functional maturation, and apoptosis. However, the molecular networks underlying these events in C/EBP⑀-induced differentiation remain poorly understood. To reveal these mechanisms, we performed a detailed molecular analysis of C/EBP⑀-induced differentiation using an inducible form of C/EBP⑀. The activation of C/EBP⑀ induced growth arrest, morphological differentiation, the expression of CD11b and secondary granule proteins, and apoptosis in myeloid cell lines. Unlike C/EBP␣, C/EBP⑀ dramatically up-regulated p27 with a concomitant down-regulation of cdk4/6 and cyclin D2/A/E. Moreover, the anti-apoptotic proteins Bcl-2 and Bcl-x were down-regulated, whereas pro-apoptotic protein Bax remained unchanged. Using a variety of mutants, we revealed that these events were all regulated by the N-terminal activation domain of C/EBP⑀. Interestingly, some of the differentiation processes such as the induction of secondary granule protein genes were clearly inhibited by c-Myc; however, inhibition of apoptosis by Bcl-x did not affect the entire differentiation processes. These data indicate the N terminus of C/EBP⑀ to be solely responsible for most aspects of myeloid differentiation, and these events were differentially affected by c-Myc. CCAAT/enhancer-binding protein ⑀ (C/EBP⑀) 3 is a member of the C/EBP family of transcription factors that plays a critical role in late granulopoiesis (1, 2). The loss of C/EBP⑀ in mice leads to the terminal differentiation failure of granulocytes and eosinophils (3). Neutrophils in C/EBP⑀-deficient mice display an abnormal morphology such as hyposegmentation of the nuclei. These mice tend to die from opportunistic infections caused by the defective neutrophil func-* This work was supported in part by grants from the Ministry of Education, Science, Sports and Culture of Japan and the Naito Foundation.
Identification of a novel enhancer of CEBPE essential for granulocytic differentiation
Blood, 2019
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...
Molecular and cellular biology, 1998
The transcription factor CCAAT/enhancer binding protein alpha (C/EBPalpha) regulates a number of myeloid cell-specific genes. To delineate the role of C/EBPalpha in human granulopoiesis, we studied its expression and function in human primary cells and bipotential (granulocytic/monocytic) myeloid cell lines. We show that the expression of C/EBPalpha initiates with the commitment of multipotential precursors to the myeloid lineage, is specifically upregulated during granulocytic differentiation, and is rapidly downregulated during the alternative monocytic pathway. Conditional expression of C/EBPalpha alone in stably transfected bipotential cells triggers neutrophilic differentiation, concomitant with upregulation of the granulocyte-specific granulocyte colony-stimulating factor receptor and secondary granule protein genes. Moreover, induced expression of C/EBPalpha in bipotential precursors blocks their monocytic differentiation program. These results indicate that C/EBPalpha serves...
Molecular and Cellular Biology, 1998
The transcription factor CCAAT/enhancer binding protein α (C/EBPα) regulates a number of myeloid cell-specific genes. To delineate the role of C/EBPα in human granulopoiesis, we studied its expression and function in human primary cells and bipotential (granulocytic/monocytic) myeloid cell lines. We show that the expression of C/EBPα initiates with the commitment of multipotential precursors to the myeloid lineage, is specifically upregulated during granulocytic differentiation, and is rapidly downregulated during the alternative monocytic pathway. Conditional expression of C/EBPα alone in stably transfected bipotential cells triggers neutrophilic differentiation, concomitant with upregulation of the granulocyte-specific granulocyte colony-stimulating factor receptor and secondary granule protein genes. Moreover, induced expression of C/EBPα in bipotential precursors blocks their monocytic differentiation program. These results indicate that C/EBPα serves as a myeloid differentiatio...
Expression of C/EBPβ from the C/ebpα gene locus is sufficient for normal hematopoiesis in vivo
Blood, 2002
CCAAT/enhancer-binding proteins (C/EBPs) are critical transcriptional regulators of differentiation of hematopoietic cells. Previous studies have shown that targeted disruption of theC/ebpα gene results in a lack of granulocytic differentiation with an arrest at the stage of immature myeloblasts. By using a gene replacement strategy in which C/EBPβ was expressed from the C/ebpα gene locus of C/EBPα-null mice, we have evaluated the ability of C/EBPβ to function for C/EBPα in directing differentiation along the granulocytic pathway. We show that the morphology and the differential cell counts of the bone marrow and peripheral blood cells from C/EBPβ knockin mice are indistinguishable from those of their wild-type littermates, indicating that hematopoiesis occurs normally in these animals. Additionally, we analyzed expression of 21 myeloid-specific genes, including markers for distinct stages of granulocytic differentiation, and found no significant differences in their levels of expre...
Acetylation of C/EBPα inhibits its granulopoietic function
Nature Communications, 2016
CCAAT/enhancer-binding protein alpha (C/EBPa) is an essential transcription factor for myeloid lineage commitment. Here we demonstrate that acetylation of C/EBPa at lysine residues K298 and K302, mediated at least in part by general control non-derepressible 5 (GCN5), impairs C/EBPa DNA-binding ability and modulates C/EBPa transcriptional activity. Acetylated C/EBPa is enriched in human myeloid leukaemia cell lines and acute myeloid leukaemia (AML) samples, and downregulated upon granulocyte-colony stimulating factor (G-CSF)-mediated granulocytic differentiation of 32Dcl3 cells. C/EBPa mutants that mimic acetylation failed to induce granulocytic differentiation in C/EBPa-dependent assays, in both cell lines and in primary hematopoietic cells. Our data uncover GCN5 as a negative regulator of C/EBPa and demonstrate the importance of C/EBPa acetylation in myeloid differentiation.
Journal of immunology (Baltimore, Md. : 1950), 2015
Neutrophil-specific granule deficiency (SGD) is a rare autosomal recessive primary immunodeficiency characterized by neutrophil dysfunction, bilobed neutrophil nuclei and lack of neutrophil-specific granules. Defects in a myeloid-specific transcription factor, CCAAT/enhancer binding protein-ε (C/EBPε), have been identified in two cases in which homozygous frameshift mutations led to loss of the leucine zipper domain. In this study, we report a 55-y-old woman affected with SGD caused by a novel homozygous 2-aa deletion (ΔRS) in the leucine zipper domain of the C/EBPε gene. The patient showed characteristic neutrophil abnormalities and recurrent skin infections; however, there was no history of deep organ infections. Biochemical analysis revealed that, in contrast to the two frameshift mutations, the ΔRS mutant maintained normal cellular localization, DNA-binding activity, and dimerization, and all three mutants exhibited marked reduction in transcriptional activity. The ΔRS mutant wa...