Regulation and function of acid sphingomyelinase (asm)/ ceramide pathway in irradiation-induced cell death

Abstract

Treating malignant glioblastoma with radiation therapy in combination with chemotherapy is one of the most promising outcomes. The present study aimed to elucidate the underlying mechanism of IR induced activation of ASM and production of ceramide in glioblastoma. We focused on the initial event that involves the activation of ASM and production of ceramide upon irradiation in human glioblastoma cells. Irradiation of human glioblastoma cells resulted in rapid activation and translocation of ASM to plasma membrane which resulted in clustering of ceramide and formation of ceramide-enriched membrane platforms. We addressed the mechanism of ASM activation which was found to be dependent on reactive oxygen species (ROS) production. Ceramide enriched-membrane platforms are known to actively cluster signaling molecule including death receptors CD95 and CD40 (Cremesti et al 2001, Grassmé et al 2001a, Grassmé et al 2001b, Grassmé et al 2002a) or subunits of NADPH oxidase like gp91phox and p47phox (Bao et al 2010, Zhang et al 2007). Similar to these observations we found that upon irradiation gp91phox clustered in ceramide enriched-membrane platforms and further using gp91phox siRNA, inhibition in IR-induced activation of ASM was seen. This result relates to the finding from Zhang and colleagues showing NADPH oxidase-derived ROS regulate Asminitiated redox signaling in a positive feedback manner (Zhang et al 2008). Lastly we identified various interacting partners of ASM, ceramide and lipid rafts using LCMS/ MS technique. Flotillin-1 and caveolin-1 proteins were identified in irradiated samples which suggest that upon irradiation, ASM translocated to plasma membrane leads to generation of ceramide and further co-localization of ASM and ceramide in lipid raft platforms takes place. Various other proteins were identified in irradiated samples of lipid rafts including dysferlin and sortilin whose role in ASM trafficking from lysosome to plasma membrane has already been described earlier upon different stimuli other than radiation (Han et al 2012, Jin et al 2008b).