Method to Measure Sphingomyelin Synthase Activity Changes in Response to CD95L.

Sphingomyelin synthases 1 and 2 convert the anti-oncometabolite ceramide to sphingomyelin, the most abundant sphingolipid in plasma membrane. CD95L-induced ceramide increase is associated with the caspase-dependent inhibition of sphingomyelin synthesis, which enhances the mitochondrial route to apoptosis. Knocking down sphingomyelin synthase 1 or inhibiting sphingomyelin synthesis facilitates ceramide accumulation, cytochrome c release from mitochondria, and caspase-9 activation in cancer cell upon CD95L treatment. Here, we describe a method to monitor in situ sphingomyelin synthase activity changes triggered by CD95L.