Cutting Edge: RIPK1 Kinase Inactive Mice Are Viable and Protected from TNF-Induced Necroptosis In Vivo

Polykratis, Apostolos; Hermance, Nicole; Zelic, Matija; Roderick, Justine E.; Kim, Chun; Van, Trieu-My; Lee, Thomas H.; Chan, Francis Ka Ming; Pasparakis, Manolis; Kelliher, Michelle A.

doi:10.4049/jimmunol.1400590

Cited by 281 publications

(250 citation statements)

References 17 publications

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“…Nec-1s and zVAD.fmk or the RIPK1 D138N knock-in kinase-dead mutation (Polykratis et al 2014), indicating a role of RIPK1 kinase activity in the activation of caspase-8 ( Fig. 2F; Supplemental Fig.…”

Section: Spata2 Promotes the Formation Of Complex Iibmentioning

confidence: 99%

SPATA2 regulates the activation of RIPK1 by modulating linear ubiquitination

Wei

Liu

et al. 2017

Genes Dev.

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Stimulation of cells with TNFα leads to the formation of the TNF-R1 signaling complex (TNF-RSC) to mediate downstream cellular fate decision. Activation of the TNF-RSC is modulated by different types of ubiquitination and may lead to cell death, including apoptosis and necroptosis, in both RIPK1-dependent and RIPK1-independent manners. Spata2 (spermatogenesis-associated 2) is an adaptor protein recruited into the TNF-RSC to modulate the interaction between the linear ubiquitin chain assembly complex (LUBAC) and the deubiquitinase CYLD (cylindromatosis). However, the mechanism by which Spata2 regulates the activation of RIPK1 is unclear. Here, we report that Spata2-deficient cells show resistance to RIPK1-dependent apoptosis and necroptosis and are also partially protected against RIPK1-independent apoptosis. Spata2 deficiency promotes M1 ubiquitination of RIPK1 to inhibit RIPK1 kinase activity. Furthermore, we provide biochemical evidence for the USP domain of CYLD and the PUB domain of the SPATA2 complex preferentially deubiquitinating the M1 ubiquitin chain in vitro. Spata2 deficiency also promotes the activation of MKK4 and JNK and cytokine production independently of RIPK1 kinase activity. Spata2 deficiency sensitizes mice to systemic inflammatory response syndrome (SIRS) induced by TNFα, which can be suppressed by RIPK1 inhibitor Nec-1s. Thus, Spata2 can regulate inflammatory response and cell death in both RIPK1-dependent and RIPK1-independent manners.

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Section: Spata2 Promotes the Formation Of Complex Iibmentioning

confidence: 99%

SPATA2 regulates the activation of RIPK1 by modulating linear ubiquitination

Wei

Liu

et al. 2017

Genes Dev.

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“…42,43 However, in vivo deficiency of RIPK1 kinase activity is shown to be deleterious for the host after Vaccinia virus infection. 40 Our results highlight the impact of the kinase domain of RIPK1 in two models of inflammation. RIPK1 K45A mice display enhanced survival against endotoxin shock, yet they are highly susceptibility to a challenge with ST.…”

Section: Ripk1 Interacts With Various Proteins Such As Ciaps Andmentioning

confidence: 89%

“…Furthermore, the cell survival after cIAP depletion is dependent on RIPK1 kinase activity and requires TNF-R signaling, similarly to previous reports. 40 Recently, in a mouse model of inflammation in the skin that is induced by SHARPIN-deficiency, RIPK1 kinase-dead mice displayed significant rescue, 20 whereas in the models of gut inflammation the kinase activity of RIPK1 did not promote severe colitis. 24,41 Further, it was shown that RIPK1 and RIPK3 are found in a complex with caspase 8 and caspase 1 where the kinase activity of RIPK1 is dispensable.…”

Section: Ripk1 Interacts With Various Proteins Such As Ciaps Andmentioning

confidence: 99%

K45A mutation of RIPK1 results in poor necroptosis and cytokine signaling in macrophages, which impacts inflammatory responses in vivo

et al. 2016

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Receptor interacting protein kinase 1 (RIPK1) participates in several cell signaling complexes that promote cell activation and cell death. Stimulation of RIPK1 in the absence of caspase signaling induces regulated necrosis (necroptosis), which promotes an inflammatory response. Understanding of the mechanisms through which RIPK1 promotes inflammation has been unclear. Herein we have evaluated the impact of a K45A mutation of RIPK1 on necroptosis of macrophages and the activation of inflammatory response. We show that K45A mutation of RIPK1 results in attenuated necroptosis of macrophages in response to stimulation with LPS, TNFα and IFNβ in the absence of caspase signaling. Impairment in necroptosis correlated with poor phosphorylation of RIPK1, RIPK3 and reduced trimerization of MLKL. Furthermore, K45A mutation of RIPK1 resulted in poor STAT1 phosphorylation (at S727) and expression of RANTES and MIP-1α following TNF-R engagement in the absence of caspase activation. Our results further indicate that in the absence of stimulation by pathogen-associated molecular patterns (PAMPs), cellular inhibitors of apoptotic proteins (cIAPs) prevent the K45-dependent auto-phosphorylation of RIPK1, leading to resistance against necroptosis. Finally, RIPK1K45A mice displayed attenuated inflammatory response in vivo as they were significantly resistant against endotoxin shock, but highly susceptible against a challenge with Salmonella typhimurium. This correlated with reduced expression of IL-1β and ROS, and poor processing of caspase 8 by RIPK1K45A macrophages. Overall, these results indicate that K45 mediated kinase activity of RIPK1 is not only important for necroptosis but it also has a key role in promoting cytokine signaling and host response to inflammatory stimuli. Apoptosis is considered as the predominant, programmed pathway of cell death; however, recently several pathways of regulated cell death have been shown to operate in cells that are considered highly inflammatory.1,2 Although apoptosis has a major role during fetal development, 3 regulated necrotic cell death does not appear to influence fetal development. 4 Interestingly, receptor interacting protein kinase 1 (RIPK1) deficiency cause embryonic lethality, suggesting a key role of RIPK1 in host survival. 5 Besides the kinase activity, RIPK1 appears to have a scaffolding function, which may influence immune homeostasis. 6,7 Depending on the interacting partners and posttranslational modifications, RIPK1 has a multifaceted role in cell signaling and cell survival.8 Following TNF-R1 signaling, RIPK1 transitions between pro-survival and pro-cell death signaling complexes. 9,10 Necroptosis is a form of regulated necrosis of cells that operates in the absence of caspase activity 9 and is initiated by the engagement of various TLRs or cytokine receptors.11 The first cardinal signaling step in necrosome signaling is the phosphorylation of RIPK1, which leads to RIPK1-RIPK3 interaction and phosphorylation of RIPK3.12 Although necroptotic signaling has been shown to be...

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“…How RIPK1 regulates RIPK3 activity is unclear; however, steady-state hematopoiesis appears normal in kinase-inactive Ripk1 D138N/D138N mice (37), suggesting that the kinase activity of RIPK1 is not required to regulate RIPK3 activity. Moreover, genetic rescue appears specific to the hematopoietic and skin lineages (26,38), because RIPK3 deficiency has marginal effects on the overall survival of RIPK1-deficient mice (24)(25)(26).…”

Section: Discussionmentioning

confidence: 99%

Hematopoietic RIPK1 deficiency results in bone marrow failure caused by apoptosis and RIPK3-mediated necroptosis

Roderick

Hermance

Zelic

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Receptor-interacting serine/threonine-protein kinase 1 (RIPK1) is recruited to the TNF receptor 1 to mediate proinflammatory signaling and to regulate TNF-induced cell death. RIPK1 deficiency results in postnatal lethality, but precisely why Ripk1 −/− mice die remains unclear. To identify the lineages and cell types that depend on RIPK1 for survival, we generated conditional Ripk1 mice. Tamoxifen administration to adult RosaCreER T2 Ripk1fl/fl mice results in lethality caused by cell death in the intestinal and hematopoietic lineages. Similarly, Ripk1 deletion in cells of the hematopoietic lineage stimulates proinflammatory cytokine and chemokine production and hematopoietic cell death, resulting in bone marrow failure. The cell death reflected cell-intrinsic survival roles for RIPK1 in hematopoietic stem and progenitor cells, because Ripk3−/− progenitors remain TNF sensitive in vitro and fail to repopulate irradiated mice. These genetic studies reveal that hematopoietic RIPK1 deficiency triggers both apoptotic and necroptotic death that is partially prevented by RIPK3 deficiency. Therefore, RIPK1 regulates hematopoiesis and prevents inflammation by suppressing RIPK3 activation.tumor necrosis factor | programmed necrosis | hematopoietic failure T he proinflammatory cytokine TNF stimulates receptorinteracting serine/threonine-protein kinase 1 (RIPK1) ubiquitination, NFκB and MAPK activation, and induction of apoptosis or necroptosis (1, 2). TNF signaling via TNF receptor 1 (TNFR1) is highly regulated and results in the recruitment of several adapter proteins including TNFR1-associated death domain (TRADD) protein, the E3 ubiquitin ligases cellular inhibitor of apoptosis protein-1 and -2 (cIAP1/2), and TNFRassociated factor 2 (TRAF2) or 5, and the serine threonine death domain-containing kinase RIPK1 (complex I) (1). We have demonstrated that the kinase activity of RIPK1 is not required for NFκB activation (3); rather, RIPK1 is modified by the addition of Lys63-linked and linear polyubiquitin chains (3-6). Polyubiquitinated RIPK1 then recruits NEMO/IκB kinase-γ (IKKγ) to mediate IKK activation and TAK1/TAB2/3 to mediate MAPK activation, resulting in antiapoptotic and proinflammatory gene expression (7,8). Deubiquitination of RIPK1 by cylindromatosis (CYLD) results in the formation of a cytosolic complex containing TRADD, Fas-associated death domain protein (FADD), caspase-8, and RIPK1 (complex IIa) (2). Caspase-8 cleaves and inactivates RIPK1 and CYLD and stimulates apoptosis (9-11). In the absence of caspase-8 or the presence of caspase inhibitors, TNF family members and potentially other ligands stimulate the kinase activity of RIPK1 to induce necroptosis (9, 11-16). RIPK1 also is recruited to the Toll-like receptor adapter TRIF via the Rip homotypic interaction motif (RHIM) to mediate NFκB activation (17) and, under conditions of caspase-8 inhibition, initiates necroptosis (14, 16). Necrostatin-1 (Nec-1), an allosteric RIPK1 inhibitor, inhibits necroptosis induced by TNF or the TLR3 ligand poly I:C and abolish...

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Cutting Edge: RIPK1 Kinase Inactive Mice Are Viable and Protected from TNF-Induced Necroptosis In Vivo

Cited by 281 publications

References 17 publications

SPATA2 regulates the activation of RIPK1 by modulating linear ubiquitination

SPATA2 regulates the activation of RIPK1 by modulating linear ubiquitination

K45A mutation of RIPK1 results in poor necroptosis and cytokine signaling in macrophages, which impacts inflammatory responses in vivo

Hematopoietic RIPK1 deficiency results in bone marrow failure caused by apoptosis and RIPK3-mediated necroptosis

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