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Section on Molecular Endocrinology, Endocrinology and Reproduction Research Branch, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, 20892
Address all correspondence and requests for reprints to: Dr. Maria L. Dufau, Building 49, Room 6A-36, 49 Covent Drive, MSC 4510, NIH 20892-4510. E-mail: dufau{at}helix.nih.gov
Our previous studies demonstrated regulation of the human LH receptor (hLHR) promoter by nuclear orphan receptors EAR2, EAR3/COUP-TFI (repression), and TR4 (activation) through a direct-repeat motif (hDR). The current studies investigated the differential binding of orphan receptors to rat (rLHR) and hLHR promoters, and their modulation of rLHR gene transcription in rat granulosa cells. The rLHR DR with one nucleotide difference from hDR at its core sequence mediated inhibition of the rLHR transcription, to which EAR2 and EAR3/COUP-TFI but not TR4 bound. The A/C mismatch was responsible for the lack of TR4 binding and function, but had no effect on EAR2 and EAR3/COUP-TFI. EAR2 and EAR3/COUP-TF bound to the rLHR DR with lower affinity than to the hDR, and exhibited lesser inhibitory capacity. This difference resulted from the lack of a guanine in the rDR, which is present 3' next to the hDR core. These studies have identified sequence-specific requirements for the binding of EAR2, EAR3/COUP-TFI, and TR4 to the DRs that explain their differential regulation of rat and human LHR genes. In addition, hCG treatment significantly reduced the inhibition of rLHR gene in granulosa cells and also decreased EAR2 and EAR3/COUP-TFI protein levels. These results indicate that hormonally regulated expression of EAR2 and EAR3/COUP-TFI contributes to gonadotropin-induced derepression of LHR promoter activity in granulosa cells.
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