Cloning and characterization of human estrogen receptor beta promoter

Estrogen receptors beta (ERbeta) belong to the nuclear receptor superfamily of ligand-dependent transcription factors that play critical roles in regulating genes involved in a wide array of biological processes. To investigate regulation of tissue-specific expression of ERbeta, we cloned and characterized a 2.1-kilobase 5'-flanking region of the human ERbeta gene. Two major transcription start sites were identified by primer extension and rapid amplification of 5'-cDNA end. The human ERbeta proximal promoter contains both TATA box and initiator element (Inr) and is GC-rich with a GC content of 65%. An Alu repeat sequence containing an ER-dependent transcription enhancer exists between -1416 and -1703. The full-length 5'-flanking sequence of ERbeta fused to a luciferase reporter exhibited functional promoter activity in ERbeta-positive TSUPr1 cell, but not in ERbeta-negative DU145 cells. In addition, DNase I protection assays of the proximal promoter showed unique protection patterns with nuclear extracts from TSUPr1 cells and ERbeta negative HeLa cells, suggesting presence of cell-specific trans-acting factors that mediate tissue/cell-specific ERbeta expression. Serial deletion analysis revealed that a 293-bp region encompassing the TATA box and Inr element possesses basal promoter activity.