Abstract SY27-01: Cutting both ways: Why FADD, FLIP, and Caspase-8 are required for embryonic development

Abstract Caspase-8 has two opposing biological functions - it promotes cell death by triggering the extrinsic pathway of apoptosis, but also has a survival activity, as it is required for embryonic development, T lymphocyte activation, and resistance to necrosis induced by Tumor Necrosis Factor-α (TNF) and related family ligands. We found that development of caspase-8-deficient mice is completely rescued by ablation of Receptor Interacting Protein Kinase-3 (RIPK3). Adult animals lacking both caspase-8 and RIPK3 display a progressive lymphoaccumulative disease resembling that seen with defects in CD95 or CD95-ligand, and resist the lethal effects of CD95 ligation in vivo. We have found that caspase-8 prevents RIPK3-dependent necrosis without inducing apoptosis by functioning in a proteolytically active complex with FLICE-Like Inhibitory Protein Long (FLIPL), and this complex is required for the protective function. Caspase-8 homodimers and caspase-8-FLIP heterodimers are activated on the oligomerized adapter molecule, FADD. Therefore, while the FADD-caspase-8-FLIP complex protects against RIPK3-dependent necrosis, the FADD-caspase-8 complex does not, and necrosis ensues. In the absence of RIPK3, FADD-caspase-8 instead induces apoptosis. This idea has several ramifications. Knock-outs of FADD, or FLIP show identical embryonic lethality, but while the FADD KO is fully rescued by ablation of RIPK3, the FLIP KO is not. Our in vitro studies suggest that in the FLIP, RIPK3 DKO mouse, sensitive cell populations die by apoptosis rather than necrosis. Indeed, we found that unlike caspase-8 or FADD KO embryos, FLIP RIPK3 DKO embryos at e9.5 show focal regions of apoptosis. Another prediction from the in vitro studies is even more striking_that lethality in the FLIP RIPK3 DKO mouse should be rescued by ablation of FADD. We have now generated the FADD FLIP RIPK3 TKO mouse, and these animals are developmentally normal. In addition to providing strong genetic evidence for a model of FADD-caspase-8-FLIP interaction with RIPK1-RIPK3, these studies involve an effect that has not been previously seen to our knowledge_two lethal genotypes (one KO, one DKO) when combined into a TKO, result in normal development. We think that's pretty cool. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr SY27-01. doi:1538-7445.AM2012-SY27-01