Identification of an enhancer and an alternative promoter in the first intron of the alpha-fetoprotein gene.
In this study we have characterized a positive regulatory region located in the first intron of the alpha-fetoprotein (AFP) gene. We show that the enhancer activity of the region depends on a 44 bp sequence centered on a CACCC motif. The sequence is the target of the two zinc fingers transcription factors BKLF and YY1. The introduction of a mutation destroying the CACCC box impairs the binding of BKLF but improves that of YY1. Moreover, the mutated sequence behaves as a negative control element, suggesting that BKLF behaves as a positive factor and that YY1 is a negative one. We also demonstrate the existence of a novel, tissue-specific AFP mRNA isoform present in the yolk sac and fetal liver which initiates from an alternative promoter located approximately 100 bp downstream of the enhancer element. The transcriptional start site controlled by this new promoter (called P2), was mapped to 66 bp downstream of a TATA box. A putative AUG translation site in-frame with exon 2 of the classical gene was found 295 bp downstream of the transcription start site. Like the traditional AFP promoter (P1), the P2 promoter is active in the yolk sac and fetal liver. Embryonic stem cells with an AFP knock-in gene containing either the P2 promoter or deleted for it were isolated and comparative analysis of embryonic bodies derived from these cells suggests that the P2 promoter contributes to early expression of the AFP gene.
Scohy
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S
Université Libre de Bruxelles, IBMM, 12 Rue Pr. Jeener and Brachet, B-6041 Gosselies, Belgium.
Gabant
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Szpirer
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Szpirer
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D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D018387:Codon, Initiator / Q000235:genetics; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics*; D050981:Erythroid-Specific DNA-Binding Factors; D005091:Exons / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter / Q000235:genetics; D006801:Humans; D007438:Introns / Q000235:genetics*; D051741:Kruppel-Like Transcription Factors; D008099:Liver / Q000196:embryology / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009928:Organ Specificity; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis / Q000235:genetics; D012097:Repressor Proteins / Q000378:metabolism; D020218:Response Elements / Q000235:genetics; D013234:Stem Cells / Q000378:metabolism; D016385:TATA Box / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism; D015533:Transcriptional Activation / Q000235:genetics; D014162:Transfection; D014407:Tumor Cells, Cultured; D050986:YY1 Transcription Factor; D015017:Yolk Sac / Q000378:metabolism; D016335:Zinc Fingers; D000509:alpha-Fetoproteins / Q000235:genetics*
D018387:Codon, Initiator; D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; C501168:KLF3 protein, human; D051741:Kruppel-Like Transcription Factors; D012333:RNA, Messenger; D012097:Repressor Proteins; D014157:Transcription Factors; D050986:YY1 Transcription Factor; C494698:YY1 protein, human; C494699:Yy1 protein, mouse; D000509:alpha-Fetoproteins; D004247:DNA