The function of sodium iodide symporter (Na+/I- symporter, or NIS) in mammary epithelial cells is essential for the accumulation of I- in milk; the newborn's first source of I- for thyroid hormone synthesis. Furthermore increased mammary gland NIS expression has previously been shown in human breast cancer. Several hormones and factors including all-trans-retinoic acid (tRA) regulate the expression of NIS. In this study, using breast cancer cell lines, we established that tRA-responsive NIS expression is confined to estrogen receptor-alpha (ER alpha) positive cells and we investigated the role of ER alpha in the regulation of NIS expression. We showed that the suppression of endogenous ER alpha by RNA interference downregulates NIS expression in ER-of. positive mammary cells. Besides, in an ER alpha negative cell line, reintroduction of ER alpha resulted in the expression of NIS in a ligand-independent manner. We also identified a novel estrogen-responsive element in the promoter region of NIS that specifically binds ER alpha and mediates ER alpha-dependent activation of transcription. Our results indicate that unliganded ER alpha (apo-ER alpha) contributes to the regulation of NIS gene expression. (c) 2006 Elsevier Inc. All rights reserved.