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Ceramide-mediated clustering is required for CD95-DISC formation

Oncogene volume 22, pages 5457–5470 (2003)Cite this article

Abstract

Early events required for induction of apoptosis by CD95 are preassociation of CD95, the formation of the death-inducing signaling complex (DISC) and clustering of CD95 in distinct membrane domains. Here, we identify the molecular ordering of these events and show that the acid sphingomyelinase (ASM) functions upstream of the DISC to mediate CD95 clustering in ceramide-enriched membrane platforms, an event that is required for DISC formation. Experiments in ASM-deficient cells revealed that CD95 ligation, in the absence of ceramide generation, triggers <1% of full caspase 8 activation at the receptor. This event, however, is both necessary and sufficient to trigger translocation of ASM onto the outer leaflet of the plasma membrane, ASM activation and ceramide release, but insufficient for apoptosis induction. Ceramide-mediated CD95 clustering then amplifies the primary CD95 signaling and drives the second step of CD95 signaling, that is, formation of the DISC yielding 100% caspase activity and apoptosis. These studies suggest that the most parsimonious interpretation of the molecular ordering of the earliest events in CD95 signaling, at least in some cells, is: CD95 ligation → 1% of maximum caspase 8 activation → ASM translocation → ceramide generation → CD95 clustering → DISC formation → 100% of maximum caspase 8 activation → apoptosis.

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Abbreviations

DISC:

death-inducing signaling complex

ASM:

acid sphingomyelinase; PBL, peripheral blood lymphocytes

NPDA:

Niemann Pick desease type A; min, minutes

PHA:

phyohemagglutinin

TNF:

tumor necrosis factor

NGF:

nerve growth factor

IL-2:

Interleukin 2

Cy3:

cyanine 3.18

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Acknowledgements

The present study was supported by the American Cancer Society and grants from the National Institutes of Health CA 85704 to RK and CA 21765 to EG. EG was also supported by grants from the German Research Foundation (DFG) Gu-335/10-2 and the European Community (MANAD).

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Author notes

  1. Richard Kolesnick and Erich Gulbins: Both the authors share last authorship

Authors and Affiliations

  1. Department of Molecular Biology, University of Duisburg-Essen, Hufelandstrasse 55, Essen, 45122, Germany

    Heike Grassmé & Erich Gulbins

  2. Laboratory of Signal Transduction, Memorial Sloan-Kettering Cancer Center, New York, 10021, NY, USA

    Aida Cremesti & Richard Kolesnick

Authors
  1. Heike Grassmé
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  2. Aida Cremesti
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  3. Richard Kolesnick
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  4. Erich Gulbins
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Corresponding authors

Correspondence to Richard Kolesnick or Erich Gulbins.

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Grassmé, H., Cremesti, A., Kolesnick, R. et al. Ceramide-mediated clustering is required for CD95-DISC formation. Oncogene 22, 5457–5470 (2003). https://doi.org/10.1038/sj.onc.1206540

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