Expression of the SNAI2 transcriptional repressor is regulated by C-ceramide.

Ceramide synthase 6 (CerS6) is an enzyme that preferentially generates pro-apoptotic C-ceramide in the sphingolipid metabolic pathway. Reduced expression of CerS6 has been associated with apoptosis resistance and recent studies point to a role for CerS6 in epithelial mesenchymal transition (EMT). Because cells that undergo EMT are also more resistant to apoptosis, we hypothesized that reduced expression of CerS6 could induce changes that are associated with EMT. We found that shRNA-mediated knockdown of CerS6 increases expression of the EMT transcription factor SNAI2 but not SNAI1 or TWIST. Treatment with C-ceramide nanoliposomes (CNL) resulted in a preferential increase in C-ceramide and suppressed SNAI2 transcriptional activation and protein expression. The increase in C-ceramide following CNL treatment was dependent on CerS activity and occurred even when CerS6 shRNA was expressed. shRNA against CerS5, which like CerS6 preferentially generates C-ceramide, also decreased transcriptional activation of SNAI2, suggesting a role for C-ceramide rather than a specific enzyme in the regulation of this transcription factor. While loss of CerS6 has been associated with apoptosis resistance, we found that cells lacking this protein are more susceptible to the effects CNL. In summary, our study identifies SNAI2 as a novel target whose expression can be influenced by C-ceramide levels. The potential of CNL to suppress SNAI2 expression has important clinical implications, since elevated expression of this transcription factor has been associated with an aggressive phenotype or poor outcomes in several types of solid tumors.