INDUCTION OF OXIDATIVE STRESS BY OKADAIC ACID IS REQUIRED FOR ACTIVATION OF TRANSCRIPTION FACTOR NF-KAPPA-B

The widely used phosphatase 1 and 2A inhibitor okadaic acid is one of the many stimuli activating transcription factor NF-kappa B in culture d cells. Phosphorylation of I kappa B-alpha, one of NF-kappa B's inhib itory subunits, is a prerequisite for I kappa B degradation and the su bsequent liberation of transcriptionally active NF-kappa B. This obser vation suggested that the phosphorylation status of IlcB is influenced by an okadaic acid-sensitive phosphatase. In this study, we provide e vidence that the effect of okadaic acid on NF-kappa B activation is in direct and dependent on the production of reactive oxygen intermediate s rather than the inhibition of an I kappa B-alpha phosphatase. Okadai c acid was found to be a strong inducer of cellular H2O2 and superoxid e production in two distinct cell lines. The structurally unrelated ph osphatase inhibitor calyculin A also induced oxidative stress. The del ayed onset of reactive oxygen production in response to okadaic acid c orrelated with the delayed activation of NF-kappa B. Moreover, NF-kapp a B induction was optimal at the same okadaic acid concentration that caused optimal H2O2 production. Both reactive oxygen intermediates pro duction and NF-kappa B activation were inhibited by the antioxidant py rrolidine dithiocarbamate and 8-(diethylamino)octyl-3,4,5-trimethyoxyb enzoate, a Ca2+ chelator. Future experiments using phosphatase inhibit ors in intact cells must consider that the compounds can act as strong inducers of oxidative stress, which provides one explanation for thei r tumor-promoting activity.