Featured Article - March 2009
Short description: The structure of the Fas–FADD death domain complex reveals how it switches on apoptosis solely through oligomerization and clustering.Nature, doi: 10.1038/nature07606
The cell-surface receptor Fas, the cytoplasmic Fas-associated death domain protein (FADD), and caspase 8 together create a signaling platform — the death-inducing signaling complex (DISC) — which is essential for initiating the apoptosis pathway. DISC acts as a cellular switch: its assembly and clustering in response to a pro-apoptotic signal acts as the 'on' switch to trigger cell death.
The stimulus for apoptosis is binding of Fas ligand (FasL) to Fas, which leads to clustering of Fas and recruitment of FADD by death domains in the cytoplasmic tail of Fas. In turn, FADD recruits and activates caspase 8, a protease required for the execution of apoptosis. Despite the importance of this complex, there is no in-depth understanding of its assembly, partly because of the difficulty in obtaining it in the absence of a pro-apoptotic signal, when the components are not interacting.
Central to the function of the DISC is the interaction between Fas and FADD, but a combined structure has been hard to come by, partly because it only exists upon DISC oligomerization and clustering, and partly because of difficulties in solubilizing the complex. The structure of the Fas–FADD complex should help answer the question of how interacting death domains initiate the apoptosis pathway.
Scott et al. 1 have isolated and crystallized the human Fas–FADD death domain complex and solved its structure at 2.7 Å. They produced the complex by combining lysates from Escherichia coli expressing recombinant Fas death domains and FADD death domains at high concentrations, thus fostering interaction.
The resulting complex has a tetrameric arrangement of four FADD death domains bound to four Fas death domains. The conformation of the Fas death domain in the complex is different from that of a typical death domain, and when compared with the solution structure of Fas 2 , it has opened up by moving helix six and 'fusing' it with helix five to form one long helix the authors term the 'stem helix'. A new helix is also formed at the C terminus of Fas. Opening up of the Fas death domain exposes its FADD-binding site and simultaneously generates bridges between adjacent open Fas molecules.
When the authors superimposed the structure of full-length FADD onto the complex, they noticed that the FADD death domain had also altered. The C-terminal helix of FADD is shifted to avoid a steric clash with the newly formed C-terminal helix of Fas. They also spotted an overall conformational rearrangement, particularly of the position of the death effector and the death domain of FADD, which they think results in exposure of caspase-8-binding residues.
To validate their findings in vivo, Scott et al. created a mutation at position 313 in the Fas death domain that forces the domain to remain open, and so should produce a hyperactive receptor. It did indeed result in significantly higher levels of apoptosis compared to the wild type.
From this the authors have produced a model of how signaling occurs in DISC. They propose that the opening of the Fas death domain is central to Fas–Fas clustering and to recruitment of FADD. The Fas–Fas bridge and FADD binding are governed by weak protein–protein interactions, which are only stabilized upon processive clustering. So clustering acts as a regulatory switch.
This model could serve as a template for other signaling platforms that lack enzymatic components, but regulate cascades through oligomeric interactions.
F. L. Scott et al. The Fas–FADD death domain complex structure unravels signalling by receptor clustering.
B. Huang, M. Eberstadt, E. T. Olejiniczak, R. P. Meadows & S. W. Fesik NMR structure and mutagenesis of the Fas (APO-1/CD95) death domain.
Nature 384, 638-641 (1996). doi:10.1038/384638a0