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Updated: Jan 3, 2026
Measuring Composition of CD95 Death-Inducing Signaling Complex and Processing of Procaspase-8 in this Complex
Published on: August 2, 2021
Long and short isoforms of c-FLIP act as control checkpoints of DED filament assembly
Laura K Hillert1, Nikita V Ivanisenko2, Johannes Espe1
1Translational Inflammation Research, Medical Faculty, Center of Dynamic Systems, Otto von Guericke University Magdeburg, Magdeburg, 39106, Germany.
Abstract:
The assembly of the death-inducing signaling complex (DISC) and death effector domain (DED) filaments at CD95/Fas initiates extrinsic apoptosis. Procaspase-8 activation at the DED filaments is controlled by short and long c-FLIP isoforms. Despite apparent progress in understanding the assembly of CD95-activated platforms and DED filaments, the detailed molecular mechanism of c-FLIP action remains elusive. Here, we further addressed the mechanisms of c-FLIP action at the DISC using biochemical assays, quantitative mass spectrometry, and structural modeling. Our data strongly indicate that c-FLIP can bind to both FADD and procaspase-8 at the DED filament. Moreover, the constructed in silico model shows that c-FLIP proteins can lead to the formation of the DISCs comprising short DED filaments as well as serve as bridging motifs for building a cooperative DISC network, in which adjacent CD95 DISCs are connected by DED filaments. This network is based on selective interactions of FADD with both c-FLIP and procaspase-8. Hence, c-FLIP proteins at the DISC control initiation, elongation, and composition of DED filaments, playing the role of control checkpoints. These findings provide new insights into DISC and DED filament regulation and open innovative possibilities for targeting the extrinsic apoptosis pathway.
Insights
Cellular apoptosis is initiated by the death-inducing signaling complex (DISC) and death effector domain (DED) filaments. This study reveals c-FLIP proteins control DISC assembly and DED filament formation, acting as crucial checkpoints.
Area of Science:
- Cellular biology
- Molecular mechanisms of apoptosis
Background:
- Extrinsic apoptosis is initiated by the assembly of the death-inducing signaling complex (DISC) at CD95/Fas.
- Procaspase-8 activation within death effector domain (DED) filaments is regulated by c-FLIP isoforms.
- The precise molecular role of c-FLIP in DISC assembly and DED filament formation remains incompletely understood.
Purpose of the Study:
- To elucidate the detailed molecular mechanisms of c-FLIP action within the DISC.
- To investigate the interactions of c-FLIP with FADD and procaspase-8 at the DED filament.
- To understand how c-FLIP influences DED filament organization and DISC network formation.
Main Methods:
- Biochemical assays to study protein interactions.
- Quantitative mass spectrometry for proteomic analysis.
- In silico structural modeling to visualize complex formation.
Main Results:
- c-FLIP directly binds to both FADD and procaspase-8 at the DED filament.
- c-FLIP facilitates the formation of shorter DED filaments and acts as a bridge in a cooperative DISC network.
- A network model demonstrates adjacent DISCs connected by DED filaments, mediated by selective FADD interactions.
Conclusions:
- c-FLIP proteins regulate the initiation, elongation, and composition of DED filaments at the DISC.
- c-FLIP acts as a critical control checkpoint in extrinsic apoptosis.
- These findings offer new insights into DISC and DED filament regulation, with potential for targeting apoptosis pathways.
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