Phosphatidylinositol 3-Kinase, Not Extracellular Signal-regulated Kinase, Regulates Activation of the Antioxidant-Responsive Element in IMR-32 Human Neuroblastoma Cells

The antioxidant-responsive element (ARE) plays an important role in the induction of phase II detoxifying enzymes including NADPH:quinone oxidoreductase (NQO1). We report herein that activation of the human NQO1-ARE (hNQO1-ARE) by tert-butylhydroquinone (tBHQ) is mediated by phosphatidylinositol 3-kinase (PI3-kinase), not extracellular signal-regulated kinase (Erk1/2), in IMR-32 human neuroblastoma cells. Treatment with tBHQ significantly increased NQO1 protein without activation of Erk1/2. In addition, PD 98059 (a selective mitogen-activated kinase/Erk kinase inhibitor) did not inhibit hNQO1-ARE-luciferase expression or NQO1 protein induction by tBHQ. Pretreatment with LY 294002 (a selective PI3-kinase inhibitor), however, inhibited both hNQO1-ARE-luciferase expression and endogenous NQO1 protein induction. In support of a role for PI3-kinase in ARE activation we show that: 1) transfection of IMR-32 cells with constitutively active PI3-kinase selectively activated the ARE in a dose-dependent manner that was completely inhibited by treatment with LY 294002; 2) pretreatment of cells with the PI3-kinase inhibitors, LY 294002 and wortmannin, significantly decreased NF-E2-related factor 2 (Nrf2) nuclear translocation induced by tBHQ; and 3) ARE activation by constitutively active PI3-kinase was blocked completely by dominant negative Nrf2. Taken together, these data clearly show that ARE activation by tBHQ depends on PI3-kinase, which lies upstream of Nrf2.