Bisphenol A stabilizes Nrf2 via Ca²⁺ influx by direct activation of the IP₃ receptor

Bisphenol A (BPA) is an endocrine-disrupting chemical used in polycarbonate and epoxy resins. Previously, we found that BPA stabilized the protein levels of nuclear factor erythroid 2-related factor 2 (Nrf2) by inducing Ca²⁺ efflux into the cytosol, followed by nitric oxide synthase activation, resulting in the enhanced nitrosylation of Keap1, which is a negative regulator of Nrf2. However, the mechanisms behind the stimulation of Ca²⁺ efflux by BPA remain unknown. In the present study, we found that BPA stimulated Ca²⁺ efflux into the cytosol from the ER, but not from outside of cells through the plasma membrane in Hep3B cells. Ca²⁺ efflux and Nrf2 stabilization by BPA were inhibited by an inhibitor of the inositol 1,4,5-trisphosphate (IP₃) receptor, 2-aminoethoxydiphenylborane, in the endoplasmic reticulum. IP₃ is produced by activation of phospholipase C (PLC) from a membrane lipid, phosphatidylinositol 4,5-bisphosphate (PIP₂). The induction of Nrf2 by BPA was not inhibited by a PLC inhibitor, U-73122, suggesting that BPA does not induce the production of IP₃ via PLC activation. We found that BPA bound directly to the IP₃ binding core domain of the IP₃ receptor, and BPA competed with IP₃ on this site. In addition, overexpression of this domain of the IP₃ receptor in Hep3B cells inhibited the stabilization of Nrf2 by BPA. These results clarified that the IP₃ receptor is a new target of BPA, and that BPA induces Ca²⁺ efflux from the endoplasmic reticulum via activation of the IP₃ receptor.