Abstract:
The role of the sphingolipid ceramide in modulating the immune response has been controversial, in part because of conflicting data regarding its ability to regulate the transcription factor NF-κB. To help clarify this role, we investigated the effects of ceramide on IL-2, a central NF-κB target. We found that ceramide inhibited protein kinase C (PKC)-mediated activation of NF-κB. Ceramide was found to significantly reduce the kinase activity of PKCθ as well as PKCα, the critical PKC isozymes involved in TCR-induced NF-κB activation. This was followed by strong inhibition of IL-2 production in both Jurkat T leukemia and primary T cells. Exogenous sphingomyelinase, which generates ceramide at the cell membrane, also inhibited IL-2 production. As expected, the repression of NF-κB activation by ceramide led to the reduction of transcription of the IL-2 gene in a dose-dependent manner. Inhibition of IL-2 production by ceramide was partially overcome when NF-κB nuclear translocation was reconstituted with activation of a PKC-independent pathway by TNF-α or when PKCθ was overexpressed. Importantly, neither the conversion of ceramide to complex glycosphingolipids, which are known to have immunosuppressive effects, nor its hydrolysis to sphingosine, a known inhibitor of PKC, was necessary for its inhibitory activity. These results indicate that ceramide plays a negative regulatory role in the activation of NF-κB and its targets as a result of inhibition of PKC.