Faculty Opinions recommendation of NINJ1 mediates plasma membrane rupture during lytic cell death.

2021 ◽  
Author(s):  
Petr Broz
Keyword(s):  
Cell Death ◽  
Plasma Membrane ◽  
Membrane Rupture

scholarly journals The Card19 locus of murine chromosome 13 regulates terminal cell lysis downstream of caspase activation and Gasdermin-D cleavage

2021 ◽  
Author(s):  
Elisabet Bjanes ◽  
Reyna Garcia Sillas ◽  
Rina Matsuda ◽  
Benjamin Demarco ◽  
Timothée Fettrelet ◽  
...  
Keyword(s):  
Cell Death ◽  
Plasma Membrane ◽  
Cell Lysis ◽  
Terminal Cell ◽  
Snp Analysis ◽  
Caspase Activation ◽  
Chromosome 13 ◽  
Murine Chromosome ◽  
Membrane Rupture ◽  
Yersinia Infection

Cell death plays a critical role in inflammatory responses. During pyroptosis, inflammatory caspases cleave Gasdermin D (GSDMD) to release an N-terminal fragment that generates plasma membrane pores that mediate cell lysis and IL-1 cytokine release. Terminal cell lysis and IL-1β release following caspase activation can be uncoupled in certain cell types or in response to particular stimuli, a state termed hyperactivation. However, the factors and mechanisms that regulate terminal cell lysis downstream of GSDMD cleavage remain poorly understood. In the course of studies to define regulation of pyroptosis during Yersinia infection, we identified a line of Card19-/- mice whose macrophages were protected from cell death and showed reduced pore formation during apoptosis or pyroptosis, yet had wild-type levels of caspase activation, IL-1 secretion, and GSDMD cleavage. Unexpectedly, CARD19, a mitochondrial CARD-containing protein, was not directly responsible for this, as two independently-generated CRISPR/Cas9 Card19 knockout mice showed no defect in macrophage cell lysis. The original Card19-/- line was generated in a 129SvEvBrd background, and SNP analysis revealed a six megabase region of 129 origin co-segregating with the Card19 locus. Card19 is located on chromosome 13, adjacent to Ninj1, which was recently reported to regulate cell lysis downstream of GSDMD activation. Nonetheless, we could not detect major defects in NINJ1 protein expression or mutations in Ninj1 coding sequence in Card19-/- mice. Mice from the original Card19-/- line exhibited significantly increased susceptibility to Yersinia infection, demonstrating that cell lysis itself plays a key role in protection against bacterial infection. Our findings identify a locus on murine chromosome 13 that regulates the ability of macrophages to undergo plasma membrane rupture downstream of gasdermin cleavage, and implicates additional NINJ1-independent factors that control terminal cell lysis.

scholarly journals Genetic targeting of Card19 is linked to disrupted NINJ1 expression, impaired cell lysis, and increased susceptibility to Yersinia infection

2021 ◽  
Vol 17 (10) ◽  
pp. e1009967
Author(s):  
Elisabet Bjanes ◽  
Reyna Garcia Sillas ◽  
Rina Matsuda ◽  
Benjamin Demarco ◽  
Timothée Fettrelet ◽  
...  
Keyword(s):  
Cell Death ◽  
Plasma Membrane ◽  
Inflammatory Responses ◽  
Critical Role ◽  
Cell Lysis ◽  
Terminal Cell ◽  
Caspase Activation ◽  
Membrane Rupture ◽  
Genetic Targeting ◽  
Increased Susceptibility

Cell death plays a critical role in inflammatory responses. During pyroptosis, inflammatory caspases cleave Gasdermin D (GSDMD) to release an N-terminal fragment that generates plasma membrane pores that mediate cell lysis and IL-1 cytokine release. Terminal cell lysis and IL-1β release following caspase activation can be uncoupled in certain cell types or in response to particular stimuli, a state termed hyperactivation. However, the factors and mechanisms that regulate terminal cell lysis downstream of GSDMD cleavage remain poorly understood. In the course of studies to define regulation of pyroptosis during Yersinia infection, we identified a line of Card19-deficient mice (Card19lxcn) whose macrophages were protected from cell lysis and showed reduced apoptosis and pyroptosis, yet had wild-type levels of caspase activation, IL-1 secretion, and GSDMD cleavage. Unexpectedly, CARD19, a mitochondrial CARD-containing protein, was not directly responsible for this, as an independently-generated CRISPR/Cas9 Card19 knockout mouse line (Card19Null) showed no defect in macrophage cell lysis. Notably, Card19 is located on chromosome 13, immediately adjacent to Ninj1, which was recently found to regulate cell lysis downstream of GSDMD activation. RNA-seq and western blotting revealed that Card19lxcn BMDMs have significantly reduced NINJ1 expression, and reconstitution of Ninj1 in Card19lxcn immortalized BMDMs restored their ability to undergo cell lysis in response to caspase-dependent cell death stimuli. Card19lxcn mice exhibited increased susceptibility to Yersinia infection, whereas independently-generated Card19Null mice did not, demonstrating that cell lysis itself plays a key role in protection against bacterial infection, and that the increased infection susceptibility of Card19lxcn mice is attributable to loss of NINJ1. Our findings identify genetic targeting of Card19 being responsible for off-target effects on the adjacent gene Ninj1, disrupting the ability of macrophages to undergo plasma membrane rupture downstream of gasdermin cleavage and impacting host survival and bacterial control during Yersinia infection.

scholarly journals Charcot-Leyden crystal formation is closely associated with eosinophil extracellular trap cell death

Blood ◽  
2018 ◽  
Vol 132 (20) ◽  
pp. 2183-2187 ◽  
Author(s):  
Shigeharu Ueki ◽  
Takahiro Tokunaga ◽  
Rossana C. N. Melo ◽  
Hidekazu Saito ◽  
Kohei Honda ◽  
...  
Keyword(s):  
Cell Death ◽  
Plasma Membrane ◽  
Protein Crystallization ◽  
Intracellular Localization ◽  
Nicotinamide Adenine Dinucleotide Phosphate ◽  
Crystal Formation ◽  
Membrane Rupture ◽  
Extracellular Traps ◽  
Extracellular Trap ◽  
Trap Cell

Abstract Protein crystallization in human tissue rarely occurs. Charcot-Leyden crystals (CLCs) were described in various eosinophilic diseases >150 years ago, but our understanding of CLC formation still remains limited. In this study, we demonstrate that CLCs observed in varied inflamed human tissues are closely associated with eosinophil cell-free granules and nuclear envelope/plasma membrane disintegration with release of filamentous chromatin (extracellular traps), typical morphologies of a regulated pathway of extracellular trap cell death (ETosis). During the process of eosinophil ETosis, eccentrically localized cytoplasmic and perinuclear CLC protein (galectin-10) is homogeneously redistributed in the cytoplasm. Rapid (1-2 minutes) formation of intracytoplasmic CLCs was observed using time-lapse imaging. Plasma membrane rupture enabled the release of both intracellularly formed CLCs and soluble galectin-10 that further contributed to formation of CLCs extracellularly, in parallel with the expulsion of free intact granules and extracellular traps. CLC formation and galectin-10 release were dependent on nicotinamide adenine dinucleotide phosphate oxidase activation. To our knowledge, this is the first demonstration of natural formation of CLCs in association with an active physiological process (ie, ETosis). These results indicate that dynamic changes in intracellular localization and release of galectin-10 contribute to CLC formation in vivo and suggest that CLC/galectin-10 might serve as an indicator of ETosis.

scholarly journals Ferroptotic pores induce Ca2+ fluxes and ESCRT-III activation to modulate cell death kinetics

2020 ◽  
Author(s):  
Lohans Pedrera ◽  
Rafael A. Espiritu ◽  
Uris Ros ◽  
Josephine Weber ◽  
Anja Schmitt ◽  
...  
Keyword(s):  
Lipid Peroxidation ◽  
Cell Death ◽  
Plasma Membrane ◽  
Membrane Damage ◽  
Protective Mechanism ◽  
Membrane Repair ◽  
Membrane Rupture ◽  
Plasma Membrane Damage ◽  
Regulated Necrosis ◽  
Sustained Increase

AbstractFerroptosis is an iron-dependent form of regulated necrosis associated with lipid peroxidation. Despite its key role in the inflammatory outcome of ferroptosis, little is known about the molecular events leading to the disruption of the plasma membrane during this type of cell death. Here we show that a sustained increase in cytosolic Ca2+ is a hallmark of ferroptosis that precedes complete bursting of the cell. We report that plasma membrane damage leading to ferroptosis is associated with membrane nanopores of a few nanometers in radius and that ferroptosis, but not lipid peroxidation, can be delayed by osmoprotectants. Importantly, Ca2+ fluxes during ferroptosis induce the activation of the ESCRT-III-dependent membrane repair machinery, which counterbalances the kinetics of cell death and modulates the immunological signature of ferroptosis. Our findings with ferroptosis provide a unifying concept that sustained increase of cytosolic Ca2+ prior to plasma membrane rupture is a common feature of regulated types of necrosis and position ESCRT-III activation as a general protective mechanism in these lytic cell death pathways.

scholarly journals Bayonets over bombs: RIPK3 and MLKL restrict Listeria without triggering necroptosis

2019 ◽  
Vol 218 (6) ◽  
pp. 1773-1775 ◽  
Author(s):  
Ting Zhang ◽  
Siddharth Balachandran
Keyword(s):  
Cell Death ◽  
Plasma Membrane ◽  
Epithelial Cells ◽  
Necrotic Cell Death ◽  
Necrotic Cell ◽  
Membrane Rupture ◽  
Cell Biol ◽  
Bacterial Replication

RIPK3 induces necroptosis by phosphorylating MLKL, which then induces plasma membrane rupture and necrotic cell death. In this issue, Sai et al. (2019. J. Cell Biol. https://doi.org/10.1083/jcb.201810014) show that RIPK3-MLKL signaling in epithelial cells promotes Listeria clearance by directly suppressing cytosolic bacterial replication, without activating cell death.

scholarly journals LPS Mediates Bovine Endometrial Epithelial Cell Pyroptosis Directly Through Both NLRP3 Classical and Non-Classical Inflammasome Pathways

2021 ◽  
Vol 12 ◽  
Author(s):  
Xiaoyu Ma ◽  
Yajuan Li ◽  
Wenxiang Shen ◽  
Ayodele Olaolu Oladejo ◽  
Jie Yang ◽  
...  
Keyword(s):  
Cell Death ◽  
Plasma Membrane ◽  
Epithelial Cell ◽  
Cell Swelling ◽  
Endometrial Epithelial Cell ◽  
Membrane Rupture ◽  
Caspase 1 ◽  
Vector Construction ◽  
Mrna And Protein Expression

As a highly inflammatory form of programmed cell death, pyroptosis is triggered by pro-inflammatory signals and associated with inflammation. It is characterized by cell swelling and large bubbles emerging from the plasma membrane, which release cytokines during inflammation. Compared with other types of cell death, pyroptosis has a distinct morphology and mechanism and involves special inflammasome cascade pathways. However, the inflammasome mechanism through which endometrial epithelial cell pyroptosis occurs in LPS-mediated inflammation remains unclear. We confirmed that there was an increased mRNA and protein expression of the IL-6, TNF-α, IL-1β, IL-18 cytokines, the inflammasome molecules NLRP3, CASPASE-1, CASPASE-4, and GSDMD in LPS-induced primary bovine endometrial epithelial cells (BEECs) in an in vitro established inflammatory model using ELISA, real-time PCR (RT-PCR), vector construction and transfection, and Western blotting. Scanning electron microscopy and lactate dehydrogenase (LDH) activity assays revealed induced cell membrane rupture, which is the main characteristic of pyroptosis. In conclusion, the cytolytic substrate GSDMD’s cleavage by caspase-1 or caspase-4 through the NLRP3 classical and non-classical inflammasome pathways, GSDMD N-terminus bind to the plasma membrane to form pores and release IL -18, IL-1β cause cell death during LPS induced BEECs inflammation.

scholarly journals Glycine targets NINJ1-mediated plasma membrane rupture to provide cytoprotection

2021 ◽  
Author(s):  
Jazlyn P Borges ◽  
Allen Volchuk ◽  
Bridget Kilburn ◽  
Neil M Goldenberg ◽  
Benjamin Ethan Steinberg
Keyword(s):  
Cell Death ◽  
Plasma Membrane ◽  
Tissue Injury ◽  
Apoptotic Cell ◽  
Molecular Target ◽  
Tissue Preservation ◽  
Downstream Process ◽  
Membrane Rupture ◽  
Cell Death Pathways ◽  
Terminal Effector

First recognized more than 30 years ago, glycine is known to protect cells against plasma membrane rupture from diverse types of tissue injury. This robust and widely observed effect has been speculated to target a late downstream process common to multiple modes of tissue injury. The molecular target and mechanism of glycine cytoprotection, however, remain entirely elusive. We hypothesized that glycine targets ninjurin-1 (NINJ1), a newly identified executioner of plasma membrane rupture in pyroptosis, necrosis, and apoptotic cell death. This common terminal effector is thought to cluster within the plasma membrane to cause cell rupture. Here, we first demonstrate that NINJ1 knockout functionally and morphologically phenocopies glycine cytoprotection in macrophages stimulated to undergo lytic cell death. Glycine treatment in NINJ1 knockout cells provides no additional protective effect. Next, we show that glycine treatment prevents NINJ1 clustering within the plasma membrane thereby preserving its integrity. By identifying NINJ1 as a glycine target, our data help resolve the long-standing mechanism of glycine cytoprotection. This new understanding will inform the development of cell and tissue preservation strategies for pathologic conditions associated with lytic cell death pathways.

scholarly journals NINJ1 mediates plasma membrane rupture during lytic cell death

2020 ◽  
Author(s):  
Nobuhiko Kayagaki ◽  
Opher Kornfeld ◽  
Bettina Lee ◽  
Irma Stowe ◽  
Karen O'Rourke ◽  
...  
Keyword(s):  
Cell Death ◽  
Plasma Membrane ◽  
Apoptotic Cell ◽  
High Mobility ◽  
Surface Protein ◽  
Underlying Mechanism ◽  
Cell Swelling ◽  
Membrane Pore ◽  
Membrane Rupture ◽  
Ionic Fluxes

Abstract Plasma membrane rupture (PMR) is the final cataclysmic event in lytic cell death. PMR releases intracellular molecules termed damage-associated molecular patterns (DAMPs) that propagate the inflammatory response. The underlying mechanism for PMR, however, is unknown. Here we show that the ill-characterized nerve injury-induced protein 1 (NINJ1) — a cell surface protein with two transmembrane regions — plays an essential role in the induction of PMR. A forward-genetic screen of randomly mutagenized mice linked NINJ1 to PMR. Ninj1–/– macrophages exhibited impaired PMR in response to diverse inducers of pyroptotic, necrotic and apoptotic cell death, and failed to release numerous intracellular proteins including High Mobility Group Box 1 (HMGB1, a known DAMP) and Lactate Dehydrogenase (LDH, a standard measure of PMR). Ninj1–/– macrophages died, but with a distinctive and persistent ballooned morphology, attributable to defective disintegration of bubble-like herniations. Ninj1–/– mice were more susceptible than wild-type mice to Citrobacter rodentium, suggesting a role for PMR in anti-bacterial host defense. Mechanistically, NINJ1 utilized an evolutionarily conserved extracellular α-helical domain for oligomerization and subsequent PMR. The discovery of NINJ1 as a mediator of PMR overturns the long-held dogma that cell death-related PMR is a passive event. Pyroptosis is a potent inflammatory mode of lytic cell death triggered by diverse infectious and sterile insults1-3. It is driven by the pore-forming fragment of gasdermin D (GSDMD)4-7 and releases two exemplar proteins: interleukin-1β (IL-1β), a pro-inflammatory cytokine, and LDH, a standard marker of PMR and lytic cell death. An early landmark study8 predicted two sequential steps for pyroptosis: (1) initial formation of a small plasma membrane pore causing IL-1β release and non-selective ionic fluxes, and (2) subsequent PMR attributable to oncotic cell swelling. PMR releases LDH (140 kDa) and large DAMPs. While the predicted size of gasdermin pores (~18 nm inner diameter9) is large enough to release IL-1β (17 kDa, ~4.5 nm diameter), the underlying mechanism for subsequent PMR has been considered a passive osmotic lysis event.

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