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

Abstract

We previously showed that cAMP inhibits IL-1beta plus IFNgamma-induced NF-kappaB 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-kappaB was investigated. Immunofluorescent staining showed that cAMP inhibited IL-1beta plus IFNgamma-induced translocation of NF-kappaB into the nucleus. Western blot analysis showed that the IL-1beta plus IFNgamma- induced phosphorylation and degradation of IkappaBa were markedly inhibited by cAMP. Immunocomplex assay involving GST-IKK revealed that cAMP inhibited IL-1beta plus IFNgamma-induced IKK activity. The PKA inhibitors had no effect on the inhibition of NF-kappaB binding by cAMP and did not change the p65 and IKB level induced by cAMP. Overexpression of PKA increased IL-1beta plus IFNgamma-induced NF-kappaB binding. These results suggest that PKA is not essential for the inhibitory effect of cAMP on NF-kappaB binding activity in hepatocytes. We demonstrated that cAMP inhibits IL-1beta plus IFNgamma-induced NF-kappaB binding due to its blockade of the upstream signal(s) leading to IkappaB phosphorylation and degradation, and is mediated by PKA-independent signaling pathways.