Abstract
Pharmacological targeting via small molecule-based chemical probes has recently acquired an emerging importance as a valuable tool to delineate molecular mechanisms. Induction of apoptosis via CD95/Fas and TRAIL-R1/2 is triggered by the formation of the death-inducing signaling complex (DISC). Caspase-8 activation at the DISC is largely controlled by c-FLIP proteins. However molecular mechanisms of this control have just started to be uncovered. In this study we report the first-in-class chemical probe targeting c-FLIPL in the heterodimer caspase-8/c-FLIPL. This rationally designed small molecule was aimed to imitate the closed conformation of the caspase-8 L2′ loop and thereby increase caspase-8 activity after initial processing of the heterodimer. In accordance with in silico predictions, this small molecule enhanced caspase-8 activity at the DISC, CD95L/TRAIL-induced caspase activation, and subsequent apoptosis. The generated computational model provided further evidence for the proposed effects of the small molecule on the heterodimer caspase-8/c-FLIPL. In particular, the model has demonstrated that boosting caspase-8 activity by the small molecule at the early time points after DISC assembly is crucial for promoting apoptosis induction. Taken together, our study allowed to target the heterodimer caspase-8/c-FLIPL and get new insights into molecular mechanisms of its activation.
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Acknowledgements
We acknowledge the center of dynamic systems, funded by the EU-programme European Regional Development Fund, Volkswagen Foundation (VW 90315), Wilhelm Sander-Stiftung (2017.008.01), DFG (LA 2386), and Russian Science Foundation 14-44-00011 for supporting our work. We thank Dr Sabine Pietkiewicz, Claudia Arndt, and Dr Nastya Shunaeva-Zotova for the experimental help. We acknowledge Prof. Peter H. Krammer (DKFZ, Heidelberg) for providing us with anti-APO-1, C15, NF6, and 1C4 antibodies.
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LH performed experiments and contributed to the manuscript text. NI performed computational modeling and worked a lot on the manuscript text. DB, CK, and JE contributed experimentally. SP and NK contributed to the theoretical design. VI and IL designed a study and wrote a manuscript.
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Hillert, L.K., Ivanisenko, N.V., Busse, D. et al. Dissecting DISC regulation via pharmacological targeting of caspase-8/c-FLIPL heterodimer. Cell Death Differ 27, 2117–2130 (2020). https://doi.org/10.1038/s41418-020-0489-0
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DOI: https://doi.org/10.1038/s41418-020-0489-0
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