Research Papers:

Role of caspases in CD95-induced biphasic activation of acid sphingomyelinase

Mario Stephan, Bärbel Edelmann, Supandi Winoto-Morbach, Ottmar Janssen, Uwe Bertsch, Cristiana Perrotta, Stefan Schütze and Jürgen Fritsch _

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Oncotarget. 2017; 8:20067-20085. https://doi.org/10.18632/oncotarget.15379

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Mario Stephan1,*, Bärbel Edelmann2,*, Supandi Winoto-Morbach1, Ottmar Janssen1, Uwe Bertsch1, Cristiana Perrotta3, Stefan Schütze1,*, Jürgen Fritsch1,*

1Institute of Immunology, Christian-Albrechts-University of Kiel, Kiel, Germany

2Department of Hematology and Oncology, University Hospital Magdeburg, Magdeburg, Germany

3Department of Biomedical and Clinical Sciences “Luigi Sacco” (DIBIC), Università degli Studi di Milano, Milano, Italy

*These authors have contributed equally to this work

Correspondence to:

Jürgen Fritsch, email: [email protected]

Keywords: acid sphingomyelinase, ceramide, CD95 ligand, internalization, CD95-receptosomes

Received: November 16, 2016     Accepted: January 24, 2017     Published: February 16, 2017


Acid sphingomyelinase (A-SMase) plays an important role in the initiation of CD95 signaling by forming ceramide-enriched membrane domains that enable clustering and activation of the death receptors. In TNF-R1 and TRAIL-R1/R2 signaling, A-SMase also contributes to the lysosomal apoptosis pathway triggered by receptor internalization. Here, we investigated the molecular mechanism of CD95-mediated A-SMase activation, demonstrating that A-SMase is located in internalized CD95-receptosomes and is activated by the CD95/CD95L complex in a biphasic manner.

Since several caspases have been described to be involved in the activation of A-SMase, we evaluated expression levels of caspase-8, caspase-7 and caspase-3 in CD95-receptosomes. The occurrence of cleaved caspase-8 correlated with the first peak of A-SMase activity and translocation of the A-SMase to the cell surface which could be blocked by the caspase-8 inhibitor IETD.

Inhibition of CD95-internalization selectively reduced the second phase of A-SMase activity, suggesting a fusion between internalized CD95-receptosomes and an intracellular vesicular pool of A-SMase. Further analysis demonstrated that caspase-7 activity correlates with the second phase of the A-SMase activity, whereas active caspase-3 is present at early and late internalization time points. Blocking caspases-7/ -3 by DEVD reduced the second phase of A-SMase activation in CD95-receptosomes suggesting the potential role of caspase-7 or -3 for late A-SMase activation.

In summary, we describe a biphasic A-SMase activation in CD95-receptosomes indicating (I.) a caspase-8 dependent translocation of A-SMase to plasma membrane and (II.) a caspase-7 and/or -3 dependent fusion of internalized CD95-receptosomes with intracellular A-SMase-containing vesicles.

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PII: 15379