Multiple regions of p45 NF-E2 are required for beta-globin gene expression in erythroid cells.
Regulated expression of genes in the beta-globin cluster depends upon sequences located between 5 and 20 kb upstream of the epsilon gene, known as the locus control region (LCR). beta-Globin expression in murine erythroleukemia (MEL) cells depends on NF-E2, a transcription factor which binds to enhancer sequences in the LCR. To gain insight into the mechanism of globin gene activation by NF-E2, an NF-E2 null MEL cell line was used to map regions of NF-E2 required for beta-globin expression. Within the transactivation domain, two discrete proline-rich regions were required for rescue of beta-globin expression. The first was located at the N-terminus of NF-E2, while the second was located N-terminal of the cap 'n collar (CNC) domain. Other proline-rich sequences were dispensable, indicating that proline content per se does not determine NF-E2 activity. Mutations within the conserved CNC domain markedly diminished rescue of beta-globin expression. This domain was required, in addition to the basic leucine zipper domain, for DNA binding activity. The requirement for discrete proline-rich sequences within the transactivation domain suggests that globin gene expression in MEL cells depends on specific interactions between NF-E2 and downstream effector molecules.
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Department of Biochemistry, Saint Jude Children's Research Hospital, 332 North Lauderdale, Memphis, TN 38101, USA.
Ney
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D000595:Amino Acid Sequence; D000818:Animals; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D050981:Erythroid-Specific DNA-Binding Factors; D015972:Gene Expression Regulation, Neoplastic; D005914:Globins / Q000096:biosynthesis*; D004915:Leukemia, Erythroblastic, Acute; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D050983:NF-E2 Transcription Factor; D050984:NF-E2 Transcription Factor, p45 Subunit; D009687:Nuclear Proteins / Q000378:metabolism; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D011392:Proline; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017384:Sequence Deletion; D014157:Transcription Factors / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*; D015533:Transcriptional Activation; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D050983:NF-E2 Transcription Factor; D050984:NF-E2 Transcription Factor, p45 Subunit; C494693:Nfe2 protein, mouse; D009687:Nuclear Proteins; D011994:Recombinant Proteins; D014157:Transcription Factors; D005914:Globins; D011392:Proline