Ceramide Disrupts HLA Class II‐restricted Antigen Processing and Presentation

Abstract

Ceramide is a bioactive sphingolipid that mediates a variety of cell functions. However, the effect of ceramide on antigen processing and HLA class II‐mediated delivery to CD4+ T cells remains unclear. We have previously shown that intracellular ceramide levels can be regulated by ceramide anlogs that directly interfere with ceramide‐metabolizing enzymes such as acid ceramidases. In this study, we examined the actions of these cell permeable ceramide analogs on professional antigen presenting cells (APC) that constitutively express HLA class II proteins and functionally present antigens to CD4+ T cells. Our results show that the treatment of APC with ceramide analogs perturbs Ag processing and presentation in the context of HLA class II molecules. Cellular uptake of ceramide analogs into the endolysosomal compartments did not alter intracellulaer class II protein levels as confirmed by confocal microscopy and western blot analysis. Mass spectroscopic analysis showed that ceramide analogs decreased the activity of acid ceramidase elevating intracellular ceramide levels in the APC. Western blot and flow cytometric analysis confirmed that the ceramide analogs diminished acid ceramidase protein expression in APC, but not cell surface class II molecules. Data also showed that the elevation of intracellular ceramide modulates endolysosomal proteases as well as the components of the HLA class II pathway. These findings suggest that ceramide regulates multiple pathways in APC including antigen processing and presentation via HLA class II molecules.Supported by the Leukemia and Lymphoma Society (3024), American Lung Association (RG‐10435‐N), MUSC HCC Funds and NC1 PO1 CA97132