Cyclic AMP Inhibits IL-1β Plus IFNγ-Induced NF-κB Translocation in Hepatocytes by a PKA Independent Mechanism

We previously showed that cAMP inhibits IL-1β plus IFNγ-induced NF-κB binding in primary hepatocytes but the signaling mechanisms responsible for this effect are not understood. In this study, the role of PKA in mediating the effect of cAMP on NF-κB was investigated. Immunofluorescent staining showed that cAMP inhibited IL-1β plus IFNγ-induced translocation of NF-κB into the nucleus. Western blot analysis showed that the IL-1β plus IFNγ- induced phosphorylation and degradation of IκBa were markedly inhibited by cAMP. Immunocomplex assay involving GST-IKK revealed that cAMP inhibited IL-1β plus IFNγ-induced IKK activity. The PKA inhibitors had no effect on the inhibition of NF-κB binding by cAMP and did not change the p65 and IKB level induced by cAMP. Overexpression of PKA increased IL-1β plus IFNγ-induced NF-κB binding. These results suggest that PKA is not essential for the inhibitory effect of cAMP on NF-κB binding activity in hepatocytes. We demonstrated that cAMP inhibits IL-1β plus IFNγ-induced NF-κB binding due to its blockade of the upstream signal(s) leading to IκB phosphorylation and degradation, and is mediated by PKA-independent signaling pathways.