Novel intronic promoter in the rat ER alpha gene responsible for the transient transcription of a variant receptor.
To analyze the molecular origin of an ER variant, the truncated ER product-1, transiently expressed at the proestrus in lactotrope cells, we generated a 2.5-kb sequence of a genomic region upstream and downstream the specific sequence truncated ER product-1. Genomic Southern blot analysis showed that truncated ER product-1 is spliced from a noncoding leader exon localized within the intron 4 of the ER alpha gene. Analysis of the promoter sequence revealed the presence of a major transcriptional start site, a canonical TATA box and putative cis regulatory elements for pituitary specific expression as well as an E-responsive element. In transient transfection, the truncated ER product-1 promoter was transcriptionally the most active in the lactotrope cell lines (MMQ). Analysis of truncated ER product-1 functionality showed that: 1) the protein inhibited ER alpha binding to the E-responsive element in electromobility shift assays, 2) inhibited the E2 binding to ER alpha in binding assays, 3) the truncated ER product-1/ER alpha complex antagonized the transcriptional activity elicited by E2, 4) nuclear localization of green fluorescent protein-ER alpha was altered in Chinese hamster ovary cell lines stably expressing truncated ER product-1. Collectively, these data demonstrated that the protein exerts full dominant negative activity against ER alpha. Moreover, truncated ER product-1/ER alpha complex also repressed the activity of all promoters tested to date, suggesting a general inhibitory effect toward transcription. In conclusion, the data suggest that truncated ER product-1 could regulate estrogen signaling via a specific promoter in lactotrope cells.
Tiffoche
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Université de Rennes I, Interactions Cellulaires et Moléculaires, Equipe Information et Programmation Cellulaires, Centre National de la Recherche Scientifique UMR 6026, Campus de Beaulieu, Rennes Cedex 35042, France.
Vaillant
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Schausi
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Thieulant
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D000818:Animals; D001483:Base Sequence / Q000235:genetics; D002460:Cell Line; D004247:DNA / Q000378:metabolism; D004958:Estradiol / Q000378:metabolism; D047628:Estrogen Receptor alpha; D017930:Genes, Reporter / Q000502:physiology; D014644:Genetic Variation; D006801:Humans; D007438:Introns / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics*; D051381:Rats; D017208:Rats, Wistar; D011960:Receptors, Estrogen / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D012016:Reference Values; D020218:Response Elements / Q000502:physiology; D013347:Subcellular Fractions / Q000378:metabolism; D014018:Tissue Distribution; D014158:Transcription, Genetic / Q000502:physiology*
D047628:Estrogen Receptor alpha; D011960:Receptors, Estrogen; C118565:TERP-1; D004958:Estradiol; D004247:DNA