Nuclear orphan receptors regulate transcription of the gene for the human luteinizing hormone receptor.
An imperfect estrogen receptor half-site response element direct-repeat, located within the TATA-less promoter of the human luteinizing hormone receptor (hLHR), was identified as an inhibitory site for Sp1/Sp3-driven basal transcription. Isolation of proteins recognizing this site by yeast one-hybrid screening of a human placenta cDNA library revealed three nuclear orphan receptors, EAR2, EAR3/COUP-TFI, and TR4. Electrophoresis mobility shift assays demonstrated that the in vitro translated nuclear orphan receptors specifically bound the direct-repeat motif of the hLHR promoter. Also, endogenous EAR2 and EAR3/COUP-TFI from JAR cell and human testis and TR4 from testes bound this motif in electrophoresis mobility shift assays. Functional analyses in CV-1 cells showed that EAR2 and EAR3/COUP-TFI repressed the hLHR promoter activity by up to 70% in a dose-dependent and sequence-specific manner. Conversely, TR4 activated the hLHR promoter activity up to 2.5-fold through binding to the same cis-element. The stimulation was reversed by coexpression of EAR2 or EAR3/COUP-TFI, indicating their competitive binding for this site. Such recognition of a common cognate site by the proteins with antagonistic functions implies that a net regulation of the hLHR gene may result from the relative availability of repressors and activator in a physiological state. This also may contribute to the differential expression of the hLHR gene in gonadal and non-gonadal tissues.
Zhang
Y
Y
Section on Molecular Endocrinology, NICHD, National Institutes of Health, Bethesda, Maryland 20892, USA.
Dufau
M L
ML
D001483:Base Sequence; D051838:COUP Transcription Factor I; D002460:Cell Line; D018076:DNA, Complementary; D004268:DNA-Binding Proteins / Q000502:physiology*; D006801:Humans; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000502:physiology*; D011401:Promoter Regions, Genetic; D011974:Receptors, LH / Q000235:genetics*; D011987:Receptors, Steroid / Q000502:physiology*; D011988:Receptors, Thyroid Hormone; D012097:Repressor Proteins; D014157:Transcription Factors / Q000502:physiology*; D014158:Transcription, Genetic / Q000502:physiology*
D051838:COUP Transcription Factor I; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; C490515:NR2C2 protein, human; C497589:NR2F1 protein, human; C075835:NR2F6 protein, human; D009419:Nerve Tissue Proteins; C489584:Nr2f6 protein, rat; D011974:Receptors, LH; D011987:Receptors, Steroid; D011988:Receptors, Thyroid Hormone; D012097:Repressor Proteins; D014157:Transcription Factors