The atypical orphan nuclear receptor DAX-1 interacts with orphan nuclear receptor Nur77 and represses its transactivation.
DAX-1 (dosage-sensitive sex reversal adrenal hypoplasia congenital critical region on the X chromosome, gene 1) (NROB1) is an atypical member of the nuclear receptor family, which lacks the classical zinc finger DNA binding domain and acts as a coregulator of a number of nuclear receptors. In this study, we have ... found that DAX-1 is a novel coregulator of the orphan nuclear receptor Nur77 (NR4A1). We demonstrate that DAX-1 represses the Nur77 transactivation by transient transfection assays. Specific interaction between Nur77 and DAX-1 was detected by coimmunoprecipitation, yeast two-hybrid, and glutathione-S-transferase pull-down assays. The ligand binding domain of DAX-1 and the activation function-2 domain of Nur77 were determined as the direct interaction domains between DAX-1 and Nur77. In vitro competition binding assay showed that DAX-1 repressed Nur77 transactivation through the competition with steroid receptor coactivator-1 for the binding of Nur77. Moreover, DAX-1 repressed Nur77- and LH-dependent increase of cytochrome P450 protein 17 promoter activity in transient transfection assays. Furthermore, Nur77-mediated transactivation was significantly increased by down-regulation of DAX-1 expression with DAX-1 small interfering RNA in testicular Leydig cell line, K28. LH treatment induced a transient increase in Nur77 mRNA, whereas LH repressed DAX-1 expression in a time- and dose-dependent manner in K28 cells. In addition, immunohistochemical analysis showed the expression of Nur77 in mouse testicular Leydig cells. These results suggest that DAX-1 acts as a novel coregulator of the orphan nuclear receptor Nur77, and that the DAX-1 may play a key role in the regulation of Nur77-mediated steroidogenesis in testicular Leydig cells.
Mesh Terms:
Animals, Binding, Competitive, Cell Line, DAX-1 Orphan Nuclear Receptor, DNA-Binding Proteins, Gene Expression, Histone Acetyltransferases, Leydig Cells, Luteinizing Hormone, Male, Mice, Nuclear Receptor Coactivator 1, Nuclear Receptor Subfamily 4, Group A, Member 1, Promoter Regions, Genetic, Protein Binding, Receptors, Cytoplasmic and Nuclear, Receptors, Retinoic Acid, Receptors, Steroid, Repressor Proteins, Steroid 17-alpha-Hydroxylase, Transcription Factors, Transcriptional Activation
Animals, Binding, Competitive, Cell Line, DAX-1 Orphan Nuclear Receptor, DNA-Binding Proteins, Gene Expression, Histone Acetyltransferases, Leydig Cells, Luteinizing Hormone, Male, Mice, Nuclear Receptor Coactivator 1, Nuclear Receptor Subfamily 4, Group A, Member 1, Promoter Regions, Genetic, Protein Binding, Receptors, Cytoplasmic and Nuclear, Receptors, Retinoic Acid, Receptors, Steroid, Repressor Proteins, Steroid 17-alpha-Hydroxylase, Transcription Factors, Transcriptional Activation
Mol. Endocrinol.
Date: Aug. 01, 2004
PubMed ID: 15155786
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