Abstract EN:

CD95 (Fas/Apo-1)-mediated apoptosis plays a pivotal role in regulating cellular homeostasis and the elimination of infected or transformed cells. Devoid of any intrinsic enzymatic activity, CD95 transduces apoptotic death signaling through homotypic protein interactions. Actually, CD95 stimulation by its ligand CD95L induces cluster formation of CD95, called CD95-Cap, which culminates in the recruitment of the adaptor protein FADD (Fas-associated Death Domain protein). In turn, FADD aggregates caspases 8 and 10. Efficient formation of the CD95/FADD/caspases complex, called DISC (Death inducing signaling complex) induces apoptosis. In this study, we show that CD95 stimulation elicits a phospholipase Cγ1 (PLCγ1)-driven biphasic calcium mobilization through the release of endoplasmic reticulum Ca²⁺ stores and a subsequent extracellular calcium entry via the CRAC (Calcium Release-Activated Calcium) channel Orai1. We also demonstrated that suppression of this calcium signal by pharmacological or genetic approaches, promotes DISC formation, accelerates the apoptotic process and improves cell sensitivity toward CD95-mediated cell death. CD95-mediated calcium signal promotes the recruitment of protein kinase C β2 (PKCβ2) into DISC in which it transiently blocks the complex formation providing a time-of-decision period to the stimulated cell. To identify the Ca²⁺- modulated protein targets preventing DISC formation and delaying the apoptotic signal, a proteomic approach will be performed using a TAP (Tandem Affinity Purification)-Tag methodology. This study should reveal new potential therapeutic targets in order to resensitize to death tumor cells.