TRAIL receptor-2 signals apoptosis through FADD and caspase-8

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Authors: Jean-Luc Bodmer, Nils Holler, Severine Reynard, Patrizia Vinciguerra, Pascal Schneider and Peter Juo
Date: Apr. 2000
From: Nature Cell Biology(Vol. 2, Issue 4)
Publisher: Nature Publishing Group
Document Type: Report
Length: 2,481 words

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Author(s): Jean-Luc Bodmer [1]; Nils Holler [1]; Séverine Reynard [1]; Patrizia Vinciguerra [1]; Pascal Schneider [1]; Peter Juo [2]; Joe Blenis [2]; Jürg Tschopp (corresponding author) [1]

Certain cytokines of the tumour-necrosis factor (TNF) family and their cognate receptors (collectively named death receptors) are potent inducers of programmed cell death (apoptosis) [1]. One such protein is the cell-surface receptor Fas, which, upon ligand binding, trimerizes and recruits the adaptor protein FADD through the cytoplasmic death domain of Fas. FADD then binds and activates procaspase-8 (ref. 1). TRAIL, the most recently identified member of the TNF family of death ligands, can induce apoptosis in a wide variety of tumour cells but not in normal cells [2]. TRAIL induces apoptosis through two death-domain-containing receptors, TRAIL-R1 (also called death receptor (DR) 4) [3] and TRAIL-R2 (or DR5) [4, 5, 6, 7, 8, 9]. Investigation of the intracellular signalling pathways responsible for TRAIL-receptor-induced apoptosis has produced controversial results. Genetic evidence [10, 11] indicates the possible involvement of a FADD-like molecule and caspase-10 rather than of FADD itself and caspase-8. Here we characterize the signalling complex of TRAIL-R2 that is assembled in response to ligand binding. We provide evidence that FADD and caspase-8, but not caspase-10, are recruited to the receptor. Moreover, mutant cell lines that lack FADD or caspase-8 are resistant to TRAIL-induced death. Thus, TRAIL-R2 and Fas death signals rely on identical signalling molecules.

The Fas signalling pathway was elucidated by means of analysis of its death-inducing signalling complex (DISC) [12], that is, by studying the endogenous (and not overexpressed) proteins that are recruited to activated Fas. We therefore took a similar approach with respect to TRAIL signalling, using recombinant Flag-tagged soluble TRAIL (sTRAIL) and anti-Flag-antibodies to immunoprecipitate the TRAIL-receptor DISC from BJAB cells. Flag-tagged sTRAIL does not induce cell death unless it is crosslinked with anti-Flag antibodies, thereby mimicking membrane-bound TRAIL [13]. At various time points after addition of the crosslinked ligand, the TRAIL-receptor DISC was immunoprecipitated and assayed for proteins known to be involved in Fas signalling. BJAB cells express TRAIL-R1 and TRAIL-R2 complementary DNA [14], and both TRAIL-receptor proteins were incorporated into the DISC, along with FADD and caspase-8, after 5 min of stimulation (Fig. 1). Maximal recruitment of all proteins was observed after 30 min, at which time caspase-8 was converted from its precursor into the active processed form, as shown by the appearance of the p43 fragment [12] in the DISC. In contrast, the substantial amounts of caspase-10 detected in the cytoplasm were neither processed nor recruited to the DISC. Activation of caspase-8 was rapidly followed by the appearance of caspase-3 activity in the cytoplasm ( Fig. 1a).

As the TRAIL-receptor DISC isolated from BJAB cells contains a mixture of TRAIL-R1 and TRAIL-R2, we next analysed Jurkat T cells, which have been shown previously to express TRAIL-R2, but not TRAIL-R1, cDNA [14]. In agreement with this, we found that only TRAIL-R2 bound to immunoprecipitated TRAIL in Jurkat cells (Fig. 1b). Recruitment of FADD and caspase-8 to activated TRAIL-R2...

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Source Citation
Bodmer, Jean-Luc, et al. "TRAIL receptor-2 signals apoptosis through FADD and caspase-8." Nature Cell Biology, vol. 2, no. 4, Apr. 2000, pp. 241+. Accessed 23 Sept. 2021.
  

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