Ceramide inhibits IL-2 production by preventing protein kinase C-dependent NF-kappaB activation: possible role in protein kinase Ctheta regulation.

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-kappaB. To help clarify this role, we investigated the effects of ceramide on IL-2, a central NF-kappaB target. We found that ceramide inhibited protein kinase C (PKC)-mediated activation of NF-kappaB. Ceramide was found to significantly reduce the kinase activity of PKCtheta as well as PKCalpha, the critical PKC isozymes involved in TCR-induced NF-kappaB 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-kappaB 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-kappaB nuclear translocation was reconstituted with activation of a PKC-independent pathway by TNF-alpha or when PKCtheta 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-kappaB and its targets as a result of inhibition of PKC.