Abstract
The transcription factor C/EBPα is a critical mediator of myeloid differentiation and is often functionally impaired in acute myeloid leukemia. Recent studies have suggested that oncogenic FLT3 activity disrupts wild-type C/EBPα function via phosphorylation on serine 21 (S21). Despite the apparent role of pS21 as a negative regulator of C/EBPα transcription activity, the mechanism by which phosphorylation tips the balance between transcriptionally competent and inhibited forms remains unresolved. In the present study, we used immuno-affinity purification combined with quantitative mass spectrometry to delineate the proteins associated with C/EBPα on chromatin. We identified DEK, a protein with genetic links to leukemia, as a member of the C/EBPα complexes, and demonstrate that this association is disrupted by S21 phosphorylation. We confirmed that DEK is recruited specifically to chromatin with C/EBPα to enhance GCSFR3 promoter activation. In addition, we demonstrated that genetic depletion of DEK reduces the ability of C/EBPα to drive the expression of granulocytic target genes in vitro and disrupts G-CSF-mediated granulocytic differentiation of fresh human BM-derived CD34(+) cells. Our data suggest that C/EBPα and DEK coordinately activate myeloid gene expression and that S21 phosphorylation on wild-type C/EBPα mediates protein interactions that regulate the differentiation capacity of hematopoietic progenitors.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Antibodies / pharmacology
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CCAAT-Enhancer-Binding Proteins / antagonists & inhibitors
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CCAAT-Enhancer-Binding Proteins / genetics
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CCAAT-Enhancer-Binding Proteins / metabolism
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CCAAT-Enhancer-Binding Proteins / physiology*
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Cell Differentiation / drug effects
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Cell Differentiation / genetics*
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Cell Differentiation / physiology
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Cells, Cultured
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Chromosomal Proteins, Non-Histone / antagonists & inhibitors
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Chromosomal Proteins, Non-Histone / genetics
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Chromosomal Proteins, Non-Histone / metabolism
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Chromosomal Proteins, Non-Histone / physiology*
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Gene Expression Regulation / drug effects
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Gene Knockdown Techniques
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HEK293 Cells
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Hematopoiesis / drug effects
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Hematopoiesis / genetics
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Humans
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K562 Cells
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MAP Kinase Kinase Kinases / antagonists & inhibitors
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MAP Kinase Kinase Kinases / metabolism
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Myeloid Cells / drug effects
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Myeloid Cells / metabolism
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Myeloid Cells / physiology*
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Oncogene Proteins / antagonists & inhibitors
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Oncogene Proteins / genetics
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Oncogene Proteins / metabolism
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Oncogene Proteins / physiology*
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Phosphorylation / drug effects
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Phosphorylation / physiology
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Poly-ADP-Ribose Binding Proteins
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Protein Binding / drug effects
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Protein Kinase Inhibitors / pharmacology
Substances
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Antibodies
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CCAAT-Enhancer-Binding Proteins
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CEBPA protein, human
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Chromosomal Proteins, Non-Histone
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DEK protein, human
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Oncogene Proteins
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Poly-ADP-Ribose Binding Proteins
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Protein Kinase Inhibitors
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MAP Kinase Kinase Kinases