PD98059 enhanced insulin, cytokine, and growth factor activation of xanthine oxidoreductase in epithelial cells involves STAT3 and the glucocorticoid receptor

PD98059 and U0126 are organic compound inhibitors frequently used to block the activity of the MEK-1/2 protein kinase. In the present work, promoter activation analyses of xanthine oxidoreductase (XOR) in epithelial cells uncovered the unexpected opposite effect of these inhibitors on activation of XOR. Activation of an XOR-luciferase fusion gene was studied in stably transfected epithelial cells. The XOR reporter gene was activated by the epidermal growth factors (EGF), prolactin, and dexamethasone and by the acute phase cytokines (APC) IL-1, IL-6, and TNFalpha as previously reported for its native gene, and insulin further stimulated activation induced with acute phase cytokines or growth factors. Activation of the proximal promoter was blocked by inhibitors of the glucocorticoid receptor (GR), p38 MAP kinase, and U0126. Unexpectedly, PD98059 activated the promoter and significantly enhanced expression induced by insulin, APC, or growth factors. Analysis of the XOR upstream DNA and proximal promoter revealed primary roles for the GR and STAT3 in mediating the effects of PD98059 on XOR activation and protein complex formation with the promoter. STAT3 phosphotyrosine-705 was rapidly induced by PD98059, dexamethasone, and insulin. XOR activation by PD98059, dexamethasone, or insulin was superinduced by a constitutively active derivative of STAT3, while a dominant negative derivative of STAT3 blocked the enhancing effect of PD98059 on XOR activation. These data demonstrate a previously unrecognized effect of PD98059 on STAT3 and the GR that could have unanticipated consequences when used to infer the involvement of the MEK-1/2 protein kinase.