MLKL and FADD Are Critical for Suppressing Progressive Lymphoproliferative Disease and Activating the NLRP3 Inflammasome

Abstract

MLKL, a key component downstream of RIPK3, is suggested to be a terminal executor of necroptosis. Genetic studies have revealed that Ripk3 ablation rescues embryonic lethality in Fadd- or Caspase-8-deficient mice. Given that RIPK3 has also been implicated in non-necroptotic pathways including apoptosis and inflammatory signaling, it remains unclear whether the lethality in Fadd(-/-) mice is indeed caused by necropotosis. Here, we show that genetic deletion of Mlkl rescues the developmental defect in Fadd-deficient mice and that Fadd(-/-)Mlkl(-/-) mice are viable and fertile. Mlkl(-/-)Fadd(-/-) mice display significantly accelerated lymphoproliferative disease characterized by lymphadenopathy and splenomegaly when compared to Ripk3(-/-)Fadd(-/-) mice. Mlkl(-/-)Fadd(-/-) bone-marrow-derived macrophages and dendritic cells have impaired NLRP3 inflammasome activation associated with defects in ASC speck formation and NF-κB-dependent NLRP3 transcription. Our findings reveal that MLKL and FADD play critical roles in preventing lymphoproliferative disease and activating the NLRP3 inflammasome.

MLKL Activation Triggers NLRP3-Mediated Processing and Release of IL-1β Independently of Gasdermin-D

“…Our findings do not challenge the existence of these pathways; rather, they indicate that, irrespective of upstream signaling events, activation of MLKL leads to processing and activation of IL-1β. These findings may explain the recently-reported defects in NLRP3 inflammasome activation in macrophages lacking MLKL(26). Our findings indicate an inextricable link between the execution of necroptosis and the release of IL-1β, and implicate bioactive IL-1β and IL-18 as likely effectors of inflammatory responses to the necroptotic death of myeloid cells.…”

Complex Pathologic Roles of RIPK1 and RIPK3: Moving Beyond Necroptosis

“…Along these lines, even though the loss of both RIPK3 and caspase-8 alleviates many acute injuries, it leads to autoimmune disease, which is further exacerbated in Mlkl -/- ; caspase-8 -/- animals [112, 113]. Overall, we interpret these and other published data to indicate that effector mechanisms activated by RIPK1 kinase are carefully balanced and their modulation should be considered with caution as it may fundamentally change the wiring of the regulation, leading to unexpected detrimental responses.…”

“…Both groups observed that while Mlkl -/- rescued embryonic lethality due to activation of necroptosis in Caspase-8 -/- and Fadd -/- mice, these mice rapidly succumbed to extremely severe systemic autoimmune disease, lymphadenopathy, and thrombocytopenia [112, 113]. Interestingly, while the development of autoimmunity may be hypothesized to result from a deficiency of both apoptosis and necroptosis in these mice; however, cell death-deficient Ripk3 -/- ;Caspase-8 -/- and Ripk3 -/- ; Fadd -/- mice developed much milder autoimmune disease.…”

Interrogating the relevance of mitochondrial apoptosis for vertebrate development and postnatal tissue homeostasis

“…This excess cell death that destroys the vasculature of the embryo, leading to developmental arrest around E10.5 (Varfolomeev et al 1998;Yeh et al 1998Yeh et al , 2000, can be fully rescued by codeletion of RIPK3 or MLKL, the key driver and effector of necroptosis, respectively. FADD/MLKL-deficient and Caspase 8/MLKL-deficient mice, which lack literally all DR-mediated apoptosis as well as programmed necroptosis are born at a normal Mendelian frequency and show no developmental abnormalities, as do mice lacking FADD plus RIPK3 or Caspase 8 plus RIPK3 (Kaiser et al 2011;Oberst et al 2011;Alvarez-Diaz et al 2016;Zhang et al 2016). Tissue homeostasis also appears unaffected in these mice, with one notable exception, the hematopoietic compartment, which gradually expands and provokes fatal autoimmunity in aged mice (Alvarez-Diaz et al 2016;Zhang et al 2016).…”

“…FADD/MLKL-deficient and Caspase 8/MLKL-deficient mice, which lack literally all DR-mediated apoptosis as well as programmed necroptosis are born at a normal Mendelian frequency and show no developmental abnormalities, as do mice lacking FADD plus RIPK3 or Caspase 8 plus RIPK3 (Kaiser et al 2011;Oberst et al 2011;Alvarez-Diaz et al 2016;Zhang et al 2016). Tissue homeostasis also appears unaffected in these mice, with one notable exception, the hematopoietic compartment, which gradually expands and provokes fatal autoimmunity in aged mice (Alvarez-Diaz et al 2016;Zhang et al 2016). As such, all of these mutants mimic the pathology noted in lpr mutant mice that lack functional FAS/CD95 but develop normally otherwise (Watanabe-Fukunaga et al 1992).…”