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 ind uction of phase II detoxifying enzymes including NADPH:quinone oxidoreducta se (NQO1). We report herein that activation of the human NQO1-ARE (hNQO1-AR E) by tert-butylhydroquinone (tBHQ) is mediated by phosphatidylinositol 3-k inase (PI3-kinase), not extracellular signal-regulated kinase (Erk1/2), in IMR-32 human neuroblastoma cells. Treatment with tBHQ significantly increas ed NQO1 protein without activation of Erk1/2, In addition, PD 98059 (a sele ctive mitogen activated kinase/Erk kinase inhibitor) did not inhibit hNQO1- ARE-luciferase expression or NQO1 protein induction by tBHQ. Pretreatment w ith LY 294002 (a selective PI3-kinase inhibitor), however, inhibited both h NQO1-ARE-luciferase expression and endogenous NQO1 protein induction. In su pport of a role for PI3-kinase in ARE activation we show that: 1) transfect ion of IMR-32 cells with constitutively active PI3-kinase selectively activ ated the ARE in a dose-dependent manner that was completely inhibited by tr eatment with LY 294002; 2) pretreatment of cells with the PI3-kinase inhibi tors, LY 294002 and wortmannin, significantly decreased NF-Ea-related facto r 2 (Nrf2) nuclear translocation induced by tBHQ; and 3) ARE activation by constitutively active PI3-kinase was blocked completely by dominant negativ e Nrf2. Taken together, these data clearly show that ARE activation by tBHQ depends on PI3-kinase, which lies upstream of Nrf2.