title: CD95 triggers Orai1-mediated localized Ca2+ entry, regulates recruitment of protein kinase C (PKC) β2, and prevents death-inducing signaling complex formation.
abstract: The death receptor CD95 plays a pivotal role in immune surveillance and immune tolerance. Binding of CD95L to CD95 leads to recruitment of the adaptor protein Fas-associated death domain protein (FADD), which in turn aggregates caspase-8 and caspase-10. Efficient formation of the CD95/FADD/caspase complex, known as the death-inducing signaling complex (DISC), culminates in the induction of apoptosis. We show that cells exposed to CD95L undergo a reorganization of the plasma membrane in which the Ca(2+) release-activated Ca(2+) channel Orai1 and the endoplasmic reticulum-resident activator stromal interaction molecule 1 colocalize with CD95 into a micrometer-sized cluster in which the channel elicits a polarized entry of calcium. Orai1 knockdown and expression of a dominant negative construct (Orai1E106A) reveal that on CD95 engagement, the Orai1-driven localized Ca(2+) influx is fundamental to recruiting the Ca(2+)-dependent protein kinase C (PKC) β2 to the DISC. PKCβ2 in turn transiently holds the complex in an inactive status, preventing caspase activation and transmission of the apoptotic signal. This study identifies a biological role of Ca(2+) and the Orai1 channel that drives a transient negative feedback loop, introducing a lag phase in the early steps of the CD95 signal. We suggest that these localized events provide a time of decision to prevent accidental cell death.
authors: Université Rennes-1, Institut de Recherche sur la Santé, l'Environnement et le Travail/Equipe Accueil Signalisation et Réponse aux agents infectieux et Chimiques 35043 Rennes,  France.
authors: Nadine
authors: N
authors: Khadra
authors: Laurence
authors: L
authors: Bresson-Bepoldin
authors: Aubin
authors: A
authors: Penna
authors: Benjamin
authors: B
authors: Chaigne-Delalande
authors: Bruno
authors: B
authors: Ségui
authors: Thierry
authors: T
authors: Levade
authors: Anne-Marie
authors: AM
authors: Vacher
authors: Josy
authors: J
authors: Reiffers
authors: Thomas
authors: T
authors: Ducret
authors: Jean-François
authors: JF
authors: Moreau
authors: Michael D
authors: MD
authors: Cahalan
authors: Pierre
authors: P
authors: Vacher
authors: Patrick
authors: P
authors: Legembre
mesh_terms: D017209:Apoptosis / Q000502:physiology*; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism*; D053455:Caspase 10 / Q000378:metabolism; D053181:Caspase 8 / Q000378:metabolism; D002460:Cell Line; D053222:Fas Ligand Protein / Q000378:metabolism; D053200:Fas-Associated Death Domain Protein / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D047468:Immunoprecipitation; D018613:Microscopy, Confocal; D046912:Multiprotein Complexes / Q000378:metabolism*; D000071740:ORAI1 Protein; D011493:Protein Kinase C / Q000378:metabolism*; D064546:Protein Kinase C beta; D018709:Statistics, Nonparametric; D019014:fas Receptor / Q000378:metabolism*
chemical_list: D015220:Calcium Channels; D053222:Fas Ligand Protein; D053200:Fas-Associated Death Domain Protein; D046912:Multiprotein Complexes; D000071740:ORAI1 Protein; C510746:ORAI1 protein, human; D019014:fas Receptor; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D053455:Caspase 10; D053181:Caspase 8; D002118:Calcium