title: Estrogen-related receptor, hERR1, modulates estrogen receptor-mediated response of human lactoferrin gene promoter.
abstract: We have shown previously that estrogen-stimulated transcription from the human lactoferrin gene in RL95-2 endometrium carcinoma cells is mediated through an imperfect estrogen response element (ERE) at the 5 -flanking region of the gene. Upstream from the ERE, a DNA sequence (-418 to -378, FP1) was selectively protected from DNase I digestion by nuclear extracts from endometrial and mammary gland cell lines. In this report, using the electrophoresis mobility shift assay, site-directed mutagenesis, and DNA methylation interference analyses, we show that three different nuclear proteins bind to the FP1 region (C1, C2, and C3 sites). The nuclear receptor, COUP-TF, binds to the C2 site. Mutations in the C1 binding region abolish C1 complex formation and reduce estrogen-dependent transcription from the lactoferrin ERE. When the imperfect ERE of the lactoferrin gene is converted to a perfect palindromic structure, the enhancing effect of the C1 binding element for estrogen responsiveness was abolished. We isolated a complementary DNA (cDNA) clone from an RL95-2 expression library that encodes the C1 site-binding protein. The encoded polypeptide maintains 99% amino acid identity with the previously described orphan nuclear receptor hERR1. A 2.2-kilobase mRNA was detected in RL95-2 cells by the newly isolated cDNA but not by the first 180 base pair of the published hERR1 sequence. By Western analysis, a major 42-kDa protein is detected in the RL95-2 nuclear extract with antibody generated against GST-hERR1 fusion protein. Finally, we show that the hERR1 interacts with the human estrogen receptor through protein-protein contacts.
authors.affiliation: Gene Regulation Group, Laboratory of Reproductive and Developmental Toxicology, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA.
authors.forename: N
authors.initials: N
authors.lastname: Yang
authors.forename: H
authors.initials: H
authors.lastname: Shigeta
authors.forename: H
authors.initials: H
authors.lastname: Shi
authors.forename: C T
authors.initials: CT
authors.lastname: Teng
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D001940:Breast; D018269:Carcinoma, Endometrioid; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D017931:DNA Primers; D018076:DNA, Complementary; D004273:DNA, Neoplasm / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005260:Female; D015870:Gene Expression; D015723:Gene Library; D005982:Glutathione Transferase / Q000096:biosynthesis; D006801:Humans; D007781:Lactoferrin / Q000096:biosynthesis* / Q000235:genetics*; D008745:Methylation; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D018160:Receptors, Cytoplasmic and Nuclear / Q000302:isolation & purification / Q000378:metabolism*; D011960:Receptors, Estrogen / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D000097983:ERRalpha Estrogen-Related Receptor
chemical_list: D015500:Chloramphenicol O-Acetyltransferase; D017931:DNA Primers; D018076:DNA, Complementary; D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; D005982:Glutathione Transferase; D007781:Lactoferrin; D009687:Nuclear Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D011960:Receptors, Estrogen; D011994:Recombinant Proteins; D000097983:ERRalpha Estrogen-Related Receptor