Identification and Characterization of Membrane Androgen Receptors in the ZIP9 Zinc Transporter Subfamily: II. Role of Human ZIP9 in Testosterone-Induced Prostate and Breast Cancer Cell Apoptosis

Recently, we discovered a cDNA in teleost ovarian follicle cells belonging to the zinc transporter ZIP9 subfamily (SLC39A9) encoding a protein with characteristics of a membrane androgen receptor (mAR). Here, we demonstrate that human ZIP9 expressed in MDA-MB-468 breast cancer cells and stably overexpressed in human prostate cancer PC-3 cells (PC-3-ZIP9) also displays the ligand binding and signaling characteristics of a specific, high-affinity mAR. Testosterone treatment of MDA-MB-468 and PC-(3)-ZIP9 cells caused activation of G proteins and second messenger pathways as well as increases in intracellular free zinc concentrations that were accompanied by induction of apoptosis. [1,2,6,7-3H]-testosterone binding and these responses were abrogated in MDA-MB-468 cells after ZIP9 small interfering RNA (siRNA) treatment and absent in PC-3 cells transfected with empty vector, confirming that ZIP9 functions as an mAR. Testosterone treatment caused up-regulation of proapoptotic genes Bax (Bcl-2-associatedXprotein), p53 (tumor protein p53), and JNK (c-Jun N-terminal kinases) in both cell lines and increased expression of Bax, Caspase 3, and cytochrome C proteins. Treatment with a zinc chelator or a MAPK inhibitor blocked testosterone-induced increases in Bax, p53, and JNK mRNA expression. The results suggest that both androgen signaling and zinc transporter functions of ZIP9 mediate testosterone promotion of apoptosis. ZIP9 is widely expressed in human tissues and up-regulated in malignant breast and prostate tissues, suggesting that it is a potential therapeutic target for treating breast and prostate cancers. These results provide the first evidence for a mechanism mediated by a single protein through which steroid and zinc signaling pathways interact to regulate physiological functions in mammalian cells.