Induction of oxidative stress by okadaic acid is required for activation of transcription factor NF-kappa B.

Abstract The widely used phosphatase 1 and 2A inhibitor okadaic acid is one of the many stimuli activating transcription factor NF-κB in cultured cells. Phosphorylation of IκB-α, one of NF-κB's inhibitory subunits, is a prerequisite for IκB degradation and the subsequent liberation of transcriptionally active NF-κB. This observation suggested that the phosphorylation status of IκB is influenced by an okadaic acid-sensitive phosphatase. In this study, we provide evidence that the effect of okadaic acid on NF-κB activation is indirect and dependent on the production of reactive oxygen intermediates rather than the inhibition of an IκB-α 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-κB. Moreover, NF-κB induction was optimal at the same okadaic acid concentration that caused optimal H2O2 production. Both reactive oxygen intermediates production and NF-κB 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.