Abstract
The upstream Gγ-globin cAMP-response element (G-CRE) plays an important role in regulating Gγ-globin expression through binding of ATF2 and its DNA-binding partners defined in this study. ATF2 knockdown resulted in a significant reduction of γ-globin expression accompanied by decreased ATF2 binding to the G-CRE. By contrast, stable ATF2 expression in K562 cells increased γ-globin transcription which was reduced by ATF2 knockdown. Moreover, a similar effect of ATF2 on γ-globin expression was observed in primary erythroid progenitors. To understand the role of ATF2 in γ-globin expression, chromatographically purified G-CRE/ATF2-interacting proteins were subjected to mass spectrometry analysis; major binding partners included CREB1, cJun, Brg1, and histone deacetylases among others. Immunoprecipitation assays demonstrated interaction of these proteins with ATF2 and in vivo GCRE binding in CD34(+) cells undergoing erythroid differentiation which was correlated with γ-globin expression during development. These results suggest synergism between developmental stage-specific recruitments of the ATF2 protein complex and expression of γ-globin during erythropoiesis. Microarray studies in K562 cells support ATF2 plays diverse roles in hematopoiesis and chromatin remodeling.
Publication types
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Research Support, N.I.H., Extramural
MeSH terms
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Activating Transcription Factor 2 / antagonists & inhibitors
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Activating Transcription Factor 2 / genetics*
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Activating Transcription Factor 2 / metabolism
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Antigens, CD34 / genetics
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Antigens, CD34 / metabolism
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Binding Sites
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Cell Differentiation
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Cyclic AMP / metabolism
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Cyclic AMP Response Element-Binding Protein / genetics
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Cyclic AMP Response Element-Binding Protein / metabolism
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DNA Helicases / genetics
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DNA Helicases / metabolism
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Erythroid Cells / cytology
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Erythroid Cells / metabolism*
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Erythropoiesis / genetics*
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Gene Expression Profiling
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Gene Expression Regulation, Developmental*
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Histone Deacetylases / genetics
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Histone Deacetylases / metabolism
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Humans
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JNK Mitogen-Activated Protein Kinases / genetics
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JNK Mitogen-Activated Protein Kinases / metabolism
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K562 Cells
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Mass Spectrometry
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Nuclear Proteins / genetics
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Nuclear Proteins / metabolism
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Promoter Regions, Genetic
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Protein Binding
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Protein Interaction Mapping
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RNA, Messenger / genetics*
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RNA, Messenger / metabolism
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RNA, Small Interfering / genetics
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RNA, Small Interfering / metabolism
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Signal Transduction
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Transcription Factors / genetics
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Transcription Factors / metabolism
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gamma-Globins / genetics*
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gamma-Globins / metabolism
Substances
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ATF2 protein, human
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Activating Transcription Factor 2
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Antigens, CD34
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CREB1 protein, human
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Cyclic AMP Response Element-Binding Protein
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Nuclear Proteins
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RNA, Messenger
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RNA, Small Interfering
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Transcription Factors
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gamma-Globins
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Cyclic AMP
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JNK Mitogen-Activated Protein Kinases
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Histone Deacetylases
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SMARCA4 protein, human
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DNA Helicases