Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

CASP8 C362A KI MLKL KO e16.5 intestine RNAseq

ABSTRACT: Caspase-8 is a protease with both pro-death and pro-survival functions: it is required for apoptosis induced by death receptors such as TNFR1 (tumor necrosis factor receptor 1) 1, and it has a critical role in suppressing necroptosis mediated by the kinase RIPK3 (receptor interacting protein kinase 3) and the pseudokinase MLKL (mixed lineage kinase-like) 2-4. Mice lacking caspase-8 display MLKL-dependent embryonic lethality 4, as do mice expressing catalytically inactive caspase-8 mutant C362A. However, Casp8C362A/C362A Mlkl-/- mice die in the perinatal period, whereas Casp8-/- Mlkl-/- mice are viable 4, indicating that inactive caspase-8 also has a pro-death scaffolding function. Here we show that inactive caspase-8 activates pyroptosis in MLKL-deficient intestinal epithelial cells around embryonic day 18, triggering the formation of ASC specks. Accordingly, intestinal atrophy and perinatal lethality in Casp8C362A/C362A Mlkl-/- mice was prevented by loss of caspase-1. In transfection studies, inactive caspase-8 mutants C362A or C362S were found in both the triton X-100 insoluble and soluble fractions, whereas wild-type caspase-8 existed only in the soluble fraction. Moreover, inactive caspase-8 shifted co-transfected ASC into the insoluble fraction, whereas wild-type caspase-8 did not. Thus, a defense mechanism is revealed that would allow intestinal epithelial cell death in the face of pathogens expressing virulence factors to inhibit caspase-8-dependent apoptosis and necroptosis.

ORGANISM(S): Mus musculus

PROVIDER: GSE125625 | GEO | 2019/09/20

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Single cell RNA-seq of intestine from E18.5 Mlkl -/- and Casp8 C362A/C362A Mlkl -/- embryos

Project description:Caspase-8 is a protease with both pro-death and pro-survival functions: it is required for apoptosis induced by death receptors such as TNFR1 (tumour necrosis factor receptor 1), and it has a critical role in suppressing necroptosis mediated by the kinase RIPK3 (receptor interacting protein kinase 3) and the pseudokinase MLKL (mixed lineage kinase-like). Mice lacking caspase-8 display MLKL-dependent embryonic lethality, as do mice expressing catalytically inactive caspase-8 mutant C362A. However, Casp8C362A/C362A Mlkl-/- mice die in the perinatal period, whereas Casp8-/- Mlkl-/- mice are viable, indicating that inactive caspase-8 also has a pro-death scaffolding function. Here we show that caspase-8 C362A triggers ASC speck formation and caspase-1-dependent pyroptosis in MLKL-deficient intestinal epithelial cells (IECs) around embryonic day 18. Pyroptosis contributed to the perinatal lethal phenotype because a number of Casp8 C362A/C362A Mlkl-/- Casp1-/- mice survived beyond weaning. Transfection studies suggested inactive caspase-8 adopts a distinct conformation to wild-type caspase-8, enabling it to engage the caspase-1 adaptor ASC. Wild-type caspase-8 was found in the Triton X-100 soluble fraction, whereas wild-type caspase-8 inhibited with the pan-caspase inhibitor emricasan, or inactive caspase-8 mutant C362A, were detected in the insoluble fraction. Moreover, inhibited or inactive caspase-8 shifted ASC into the insoluble fraction. Perinatal lethality was recapitulated when expression of caspase-8 C362A was restricted to IECs, but intriguingly, only in the absence of MLKL. Hence, unanticipated plasticity in death pathways is revealed such that IECs can undergo caspase-1-dependent death when caspase-8-dependent apoptosis and MLKL-dependent necroptosis are inhibited.

2019-09-20 | GSE132133 | GEO

ZBP1 and TRIF trigger lethal necroptosis in mice lacking caspase-8 and TNFR1

Project description:Necroptosis is a lytic form of cell death that is mediated by the kinase RIPK3 and the pseudokinase MLKL when caspase-8 is inhibited downstream of death receptors, toll-like receptor 3 (TLR3), TLR4, and the intracellular Z-form nucleic acid sensor ZBP1. Oligomerization and activation of RIPK3 is driven by interactions with the kinase RIPK1, the TLR adaptor TRIF, or ZBP1. In this study, we use immunohistochemistry (IHC) and in situ hybridization (ISH) assays to generate a tissue atlas characterizing RIPK1, RIPK3, Mlkl, and ZBP1 expression in mouse tissues. RIPK1, RIPK3, and Mlkl were co-expressed in most immune cell populations, endothelial cells, and many mucosal epithelia. ZBP1 was expressed in many immune populations, but had more variable expression in epithelia compared to RIPK1, RIPK3, and Mlkl. Intriguingly, expression of ZBP1 was elevated in Casp8-/- Tnfr1-/- embryos prior to their succumbing to aberrant necroptosis around embryonic day 15. ZBP1 contributed to this embryonic lethality because rare Casp8-/- Tnfr1-/- Zbp1-/- mice survived until after birth. Necroptosis mediated by TRIF contributed to the demise of Casp8-/- Tnfr1-/- Zbp1-/- pups in the perinatal period. Of note, Casp8-/- Tnfr1-/- Trif-/- Zbp1-/- mice exhibited autoinflammation and morbidity, typically within 5-7 weeks of being born, which is not seen in Casp8-/- Ripk1-/- Trif-/- Zbp1-/-, Casp8-/- Ripk3-/-, or Casp8-/- Mlkl-/- mice. Therefore, after birth, loss of caspase-8 probably unleashes RIPK1-dependent necroptosis driven by death receptors other than TNFR1.

2024-04-01 | GSE254754 | GEO

Oligomerisation-driven MLKL ubiquitylation antagonises necroptosis

Project description:Mixed lineage kinase domain-like (MLKL) is the executioner in the caspase 8-independent form of programmed cell death called necroptosis. Once Receptor Interacting serine/threonine Protein Kinase 3 (RIPK3) is activated by upstream cell death signals, it phosphorylates MLKL and triggers the oligomerization and membrane translocation required for MLKL induced membrane disruption. Besides phosphorylation, MLKL also undergoes ubiquitylation during the early stages of necroptosis, yet neither the mechanism nor the significance of this event has been demonstrated. Here we show that necroptosis-specific, multi-mono-ubiquitylation of MLKL occurs on biological membranes, and requires its activation and oligomerisation. Inactive MLKL mutants recruited to membranes during necroptosis are ubiquitylated but this results in their proteasome and lysosome dependent turnover. We identified several ubiquitylated lysines however mutation of these did not affect MLKL ubiquitylation in response to a necroptotic stimulus.

2021-04-28 | PXD015537 | Pride

Caspase-8 and FADD prevent spontaneous ZBP1 expression and necroptosis

Project description:Caspase-8 and FADD play key roles in the regulation of cell death by necroptosis. The absence of either protein results in early embryonic lethality due to the activation of the kinase RIPK3 and its phosphorylation of the necroptosis executioner, MLKL. We genetically engineered and characterized a mouse model to monitor MLKL phosphorylation in the absence of necroptosis in vivo. Ablation of caspase-8 or FADD resulted in the transcriptional upregulation in several tissues of ZBP1, a cytosolic nucleic acid sensor capable of activating RIPK3, and ZBP1 was required for spontaneous phosphorylation of MLKL. Our findings provide a novel mechanism by which the FADD-Caspase-8 complex prevents necroptosis.

2022-09-19 | GSE208744 | GEO

Proteomic analysis of TRAIL-induced endogenous RIPK3-dependent necrosome complex

Project description:Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a well-known inducer of apoptosis via formation of the primary death-inducing signaling complex (TRAIL-DISC) at the level of membrane death receptors (DR4 and DR5) which recruit successively FADD and caspase-8. TRAIL can also induce necroptosis when caspases are inhibited. Necroptosis is a regulated cell death dependent on the formation of a cytosolic necrosome complex which includes RIPK1, RIPK3 and MLKL proteins. Elucidating the molecular mechanisms involved in TRAIL-induced necroptosis might provide new insights into the TRAIL death signaling pathway. Here, we report the analysis by mass spectrometry of endogenous RIPK3-dependent necrosome complex constituents upon necroptosis induced by TRAIL/z-VAD/Birinapant (TzB) in HT29 cells. Besides characterization of RIPK1, RIPK3, MLKL, FADD, caspase-8, we find TRIM21 as a new constituent of the necrosome complex. Moreover RIPK1, RIPK3, MLKL, P-MLKL, FADD, caspase-8 and TRIM21 are also found associated to the native TRAIL-DISC upon TzB stimulation showing initiation of the necrotic pathway at the level of TRAIL death receptors in HT29 cells. Finally, TRIM21 may positively modulate necroptosis induction by downregulating NF-kB activation.

2021-03-18 | PXD003383 | Pride

Human RIPK3 C-lobe phosphorylation is essential for necroptotic signaling

Project description:Necroptosis is a caspase-independent, pro-inflammatory mode of programmed cell death which relies on the activation of the terminal effector, MLKL, by the upstream protein kinase RIPK3. To mediate necroptosis, RIPK3 must stably interact with, and phosphorylate the pseudokinase domain of MLKL. While the precise molecular cue that prompts RIPK3 to activate MLKL is incompletely understood, it is known that RIPK3 is highly regulated by phosphorylation. Here, we sought to identify phosphorylation sites of RIPK3 and dissect their regulatory functions. Phosphoproteomics identified 21 phosphorylation sites in HT29 cells overexpressing human RIPK3. By comparing cells expressing wild-type and kinase-inactive D142N RIPK3, autophosphorylation sites and substrates of other cellular kinases were distinguished. Of these 21 phosphosites, extensive mutational analyses identified only pT224 and pS227 as crucial, synergistic sites for stable interaction with MLKL to promote necroptosis, while the recently reported activation loop phosphorylation at S164/T165 negatively regulate the kinase activity of RIPK3. Despite being able to phosphorylate MLKL to a similar or higher extent than the wild-type RIPK3, mutation of T224, S227, and RHIM in RIPK3 attenuated or compromised their abilities to mediate necroptosis. This finding highlights the stable recruitment of human MLKL by RIPK3 to the necrosome as an essential checkpoint in necroptosis signaling, which is independent from and precedes the phosphorylation of MLKL.

2022-07-30 | PXD033488 | Pride

Ptpn6 inhibits caspase-8- and Ripk3/Mlkl-dependent inflammation

Project description:Ptpn6 is a cytoplasmic phosphatase that functions to prevent autoimmune disease and IL-1R-dependent caspase-1-independent inflammatory disease. Conditional deletion of Ptpn6 in neutrophils (Ptpn6∆PMN) is sufficient to initiate IL-1R-dependent cutaneous inflammatory disease, but the source of IL-1 and the mechanisms behind IL-1 release remain unclear. Here, we investigated the mechanisms controlling IL-1α/β release from neutrophils by inhibiting caspase-8-dependent apoptosis and Ripk1/Ripk3/Mlkl-regulated necroptosis. Loss of Ripk1 accelerated disease onset, whereas combined deletion of caspase-8 and either Ripk3 or Mlkl strongly protected Ptpn6∆PMN mice. Ptpn6∆PMN neutrophils displayed increased p38-dependent Ripk1-independent IL-1 and TNF production, and were prone to cell death. Together, these data emphasize dual functions for Ptpn6 in the negative regulation of p38 MAP kinase activation to control TNF and IL-1α/β transcription, and in maintaining Ripk1 function to prevent caspase-8- and Ripk3/Mlkl-dependent cell death and concomitant IL-1α/β release.

2019-11-12 | GSE139223 | GEO

Ubiquitylation of RIPK3 beyond-the-RHIM limits RIPK3 activity and cell death

Project description:Pathogen recognition receptors and TNF superfamily members engage Receptor Interacting Serine/threonine Kinase-3 (RIPK3) to activate programmed cell death, including MLKL-mediated necroptosis and caspase-8-dependent apoptosis. However, the post-translational control of RIPK3 signalling is not fully understood. Using mass-spectrometry, we identified a novel ubiquitylation site on murine RIPK3 beyond the RIP homotypic interaction motif (RHIM) on K469. Complementation of RIPK3-deficient cells with a Ripk3K469R mutant demonstrated that the decoration of RIPK3 K469 by ubiquitin limits both RIPK3-mediated caspase-8 activation and apoptotic killing, in addition to RIPK3 autophosphorylation and MLKL-mediated necroptosis. Unexpectedly, the overall ubiquitylation of mutant RIPK3K469R was enhanced, which largely resulted from additional RIPK3 ubiquitylation on K359. Loss of RIPK3 K359 ubiquitylation reduced RIPK3K469R hyper-ubiquitylation and limited the ability of Ripk3K469R/K469R to trigger enhanced killing. Ripk3K469R/K469R mice challenged with Salmonella displayed increased bacterial loads in the spleen and liver, with reduced IFN serum levels. Therefore, RIPK3 K469 ubiquitylation can function to prevent RIPK3 ubiquitylation on alternate lysine residues, which otherwise promote RIPK3 oligomerization and consequent cell death signalling.

2022-08-12 | PXD026533 | Pride

ZBP1-MLKL-dependent Necroptosis Triggers STING-mediated Cytosolic DNA Sensing and Promotes Antitumor Immunity of Radiation

Project description:Ionizing radiation promotes cytosolic DNA sensing and consequent antitumor immune responses. But how tumor cell-intrinsic cytosolic DNA sensing is initiated by radiation remains poorly defined. Here, we demonstrated that STING-mediated type I interferon production in tumor cells after radiation relied on the engagement of MLKL-mediated necroptosis, which was elicited by the ZBP1-RIPK3 signaling axis. Physiologically, tumor cell-intrinsic ZBP1-MLKL cascade augmented antitumor immune responses after radiation largely by regulating STING signaling. Mechanistically, ZBP1-MLKL-dependent necroptosis maintained the enrichment of mitochondria DNA inside the cytosol of tumor cells after radiation in a cell-density dependent fashion, contributing to type I interferon responses. In contrast, ablation of caspase-8 unleashed ZBP1-MLKL cascade to gain enhanced cytosolic DNA sensing, and in turn potentiated therapeutic effects of radiation. Thus, our findings uncover an unanticipated mechanism that ZBP1-MLKL-dependent necroptosis drives cytosolic DNA sensing-mediated antitumor immunity after radiation, and provide new strategy to improve radiotherapy by inhibiting caspase-8 cascade.

2021-10-13 | GSE168016 | GEO

Human cytomegalovirus protein pUL36: a dual cell death pathway inhibitor

Project description:Human cytomegalovirus (HCMV) is an important human pathogen and a paradigm of intrinsic, innate and adaptive viral immune evasion. Here, we employ multiplexed tandem mass tag-based proteomics to characterise host proteins targeted for degradation late during HCMV infection. This approach revealed that mixed lineage kinase domain-like protein (MLKL), a key terminal mediator of cellular necroptosis, was rapidly and persistently degraded by the minimally passaged HCMV strain Merlin but not the extensively passaged strain AD169. The strain Merlin viral inhibitor of apoptosis pUL36 was necessary and sufficient both to degrade MLKL and to inhibit necroptosis. Furthermore, mutation of pUL36 Cys131 abrogated MLKL degradation and restored necroptosis. As the same residue is also required for pUL36-mediated inhibition of apoptosis by preventing proteolytic activation of pro-caspase 8, we define pUL36 as a multifunctional inhibitor of both apoptotic and necroptotic cell death.

2020-07-10 | PXD017279 | Pride

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data