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 cultured cells. Phosphorylation of I kappa B-alpha, one of NF-kappa B's inhibitory subunits, is a prerequisite for I kappa B degradation and the subsequent liberation of transcriptionally active NF-kappa B. This observation suggested that the phosphorylation status of IlcB is influenced by an okadaic acid-sensitive phosphatase. In this study, we provide evidence that the effect of okadaic acid on NF-kappa B activation is indirect and dependent on the production of reactive oxygen intermediates rather than the inhibition of an I kappa B-alpha phosphatase. Okadaic acid was found to be a strong inducer of cellular H2O2 and superoxide production in two distinct cell lines. The structurally unrelated phosphatase inhibitor calyculin A also induced oxidative stress. The delayed onset of reactive oxygen production in response to okadaic acid correlated with the delayed activation of NF-kappa B. Moreover, NF-kappa B induction was optimal at the same okadaic acid concentration that caused optimal H2O2 production. Both reactive oxygen intermediates production and NF-kappa B activation were inhibited by the antioxidant pyrrolidine dithiocarbamate and 8-(diethylamino)octyl-3,4,5-trimethyoxybenzoate, a Ca2+ chelator. Future experiments using phosphatase inhibitors in intact cells must consider that the compounds can act as strong inducers of oxidative stress, which provides one explanation for their tumor-promoting activity.