Traceless synthesis of ceramides in living cells reveals saturation-dependent apoptotic effects

Mammalian cells synthesize thousands of distinct lipids, yet the function of many of these lipid species is unknown. Ceramides, a class of sphingolipid, are implicated in several cell-signaling pathways but poor cell permeability and lack of selectivity in endogenous synthesis pathways have hampered direct study of their effects. Here we report a strategy that overcomes the inherent biological limitations of ceramide delivery by chemoselectively ligating lipid precursors in vivo to yield natural ceramides in a traceless manner. Using this method, we uncovered the apoptotic effects of several ceramide species and observed differences in their apoptotic activity based on acyl-chain saturation. Additionally, we demonstrate spatiotemporally controlled ceramide synthesis in live cells through photoinitiated lipid ligation. Our in situ lipid ligation approach addresses the long-standing problem of lipid-specific delivery and enables the direct study of unique ceramide species in live cells.