Regulation of the pro-B-cell-specific enhancer of the Id1 gene involves the C/EBP family of proteins.
The Id1 protein acts as a negative regulator in early-B-cell differentiation by antagonizing the function of the basic helix-loop-helix transcription factors. Expression of the Id1 gene during B-cell development is governed at the transcriptional level primarily by a pro-B-cell-specific enhancer (PBE) located 3 kb downstream of the gene. We report here the identification of CAAT/enhancer binding protein beta (C/EBPbeta) as a component of the two major PBE-binding complexes (PBEC1 and PBEC2) found in pro-B cells by gel mobility shift assays. Formation of the PBECs is abolished when a classic C/EBP binding site is used as a competitor, and binding complexes similar to the PBECs are formed when the classic C/EBP site is used as a probe. We show that CHOP, a negative regulator of C/EBPs, specifically inhibits PBE binding in vitro and its enhancer activity in vivo. In pro-B cells, C/EBPbeta binds to the PBE site not as apparent homodimers but possibly in association with at least one other polypeptide, which might determine the pro-B-cell-specific expression of the Id1 gene. Although isoforms of C/EBPbeta are expressed in various B cells, they bind to DNA only in LyD9 and Ba/F3 pro-B cells. We show that CHOP is expressed in 70Z/3 and WEHI-231 cells. We also demonstrate that CHOP is associated with C/EBPbeta in WEHI-231 cells, which may provide an additional mechanism to control the function of C/EBPbeta and the expression of the Id1 gene.
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Department of Cell Biology, New York University Medical Center, New York, New York 10016, USA.
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D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology / Q000502:physiology*; D022762:CCAAT-Enhancer-Binding Proteins; D002454:Cell Differentiation; D002460:Cell Line; D004247:DNA / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000032:analysis / Q000378:metabolism*; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D018507:Gene Expression Regulation, Developmental; D005796:Genes / Q000235:genetics; D051798:Inhibitor of Differentiation Protein 1; D051379:Mice; D009687:Nuclear Proteins / Q000378:metabolism*; D011485:Protein Binding; D012097:Repressor Proteins; D051742:Transcription Factor CHOP; D014157:Transcription Factors / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; C497250:Ddit3 protein, mouse; C497455:Idb1 protein, mouse; D051798:Inhibitor of Differentiation Protein 1; D009687:Nuclear Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D051742:Transcription Factor CHOP; D004247:DNA