bims-cediti 2026-06-14 papers

bims-cediti

Biomed News

on Cell death in innate immunity, inflammation, and tissue repair

Issue of 2026–06–14
twenty-six papers selected by
Kateryna Shkarina, Universität Bonn

OLT1177 (Dapansutrile) inhibits Gasdermin D-dependent IL-1β Release and Pyroptotic Cell Death in Bone Marrow-derived Macrophages.
Characterization of programmed cell death pathways activated in Mycobacterium tuberculosis-infected human macrophages.
Caspase-8 silences cell death-independent constitutive immune activation driven by tonic TNF-α.
Infection-induced glucose starvation triggers NINJ1-dependent macrophage lysis and Candida escape.
Cell size modulates ferroptosis susceptibility.
GBP1 recruitment to actin-rich pedestals of extracellular Gram-negative bacteria promotes pyroptosis.
Time-lapse confocal laser scanning microscopy analysis of FOOD formation.
Methylation of DNA duplexes regulates cGAS-mediated innate immune activation via condensate formation.
Super-resolution imaging of cell death in Drosophila tissues via expansion and pan-expansion microscopy.
Screening and identification of protein-protein interaction using proximity labeling.
Staphylococcus aureus inhibits the NLRP3 inflammasome in macrophages to varying degrees during early and late stages of infection.
Loss of erythrocyte sialic acid in sepsis disrupts inhibitory Siglec interactions, driving neutrophil hyperactivation and NET outspread.
Role of the Host Membrane Trafficking Protein Dynamin 2 in Cell-to-Cell Spread of Bacterial Pathogens.
The signaling and regulatory functions of dying cells and cell corpses.
ANXA2 promotes NLRP3 inflammasome activation and neuronal pyroptosis after intracerebral hemorrhage.
Measuring efferocytosis by intraperitoneal clearance assay.
Cell fusion after wounding requires plasma membrane damage and endocytosis.
Cell stress and death liberate the autophagy-inhibitory tissue stress hormone DBI/ACBP into the circulation.
Toll-like Receptor 3 as a Context-Dependent Molecular Switch in Epithelial Cancers: Balancing Cell Death and Tumor-Supportive Programs.
IFITM1 differentially regulates antibacterial immunity and immunopathology but is dispensable for antiparasitic responses.
Met-Vision reveals coexisting energetic states in tissue macrophages redistributed by inflammation.
Mechanistic analysis of MLKL-driven cell survival.
The role of neutrophils and neutrophil extracellular traps in Staphylococcus aureus colonization in healthy and inflamed skin.
Mechanisms driving open-closed transitions and dimer stability of the large GTPases hGBP1 and hGBP5.
Ubiquitin beyond the proteome: lipids, glycans, metabolites, nucleic acids, and an expanding molecular landscape.
STING1 senses mitochondrial damage to promote mitophagy.

bioRxiv. 2026 Jun 03. pii: 2026.05.31.729061. [Epub ahead of print]

OLT1177 (Dapansutrile) inhibits Gasdermin D-dependent IL-1β Release and Pyroptotic Cell Death in Bone Marrow-derived Macrophages.

Karl A Pankratz, Masoom Raza, Jarren Ypil, Migachelle Banks, Carlo Marchetti, Tania Azam, Charles A Dinarello, Shaikh M Atif.

Gasdermins are a family of pore-forming proteins that regulate the release of pro-inflammatory cytokine, interleukin-1β (IL-1β) from infected or PAMP-stimulated cells. During infection or injury, IL-1β is released by both human and mouse macrophages. IL-1β release from mouse macrophages is associated with cell death, often termed "pyroptosis". Mouse macrophages undergoing pyroptosis assemble an exit channel termed gasdermin D (GSDMD). Both the processing of IL-1β and the formation of the exit channel are caspase-1 dependent. Here, in bacterial endotoxin, lipopolysaccharide (LPS), treated mouse bone marrow-derived macrophages (BMDMs), we studied the pharmacologic inhibition of the intracellular nucleotide-binding domain, leucine-rich-containing family, pyrin domain- containing-3 (NLRP3) inflammasome by OLT1177. BMDMs stimulated with LPS plus the potassium efflux inducer nigericin triggered the formation of the NLRP3 inflammasome. Treatment of these BMDMs with OLT1177 suppressed cell death by 42% and ASC (apoptosis-associated speck-like protein containing a caspase recruitment domain)-speck formation by approximately 60%. In addition, OLT1177 dose-dependently inhibited IL-1β, CCL3, and myeloperoxidase (MPO) secretion and the pore-forming (GSDMD) from LPS-primed BMDMs, suggesting the existence of a vicious cycle controlled by IL-1β release. Overall, our study demonstrates that OLT1177 prevents IL-1β release from BMDMs by inhibiting caspase-1 and the conversion of (GSDMD) into its active N-terminal fragment (GSDMD-N). This study thus supports the concept that orally administered OLT1177 can be used to prevent local as well as systemic inflammation in humans.

DOI: https://doi.org/10.64898/2026.05.31.729061
Cell Death Discov. 2026 Jun 08.

Characterization of programmed cell death pathways activated in Mycobacterium tuberculosis-infected human macrophages.

Guanchao Ding, Jacques Augenstreich, Anushka Poddar, Akshaya Ganesh, Liron David, Ravin Fisher, Volker Briken.

Mycobacterium tuberculosis (Mtb) primarily infects human lung macrophages, which serve as its major replication niche. Mtb can manipulate host macrophage cell death pathways to its advantage by inhibiting apoptosis and inducing necrotic cell death. However, the specific necrotic cell death pathway activated in human macrophages after Mtb infection remains unclear. Here, we used the THP-1 cell line and primary human monocyte-derived macrophage (hMDM) to analyze multiple programmed cell death pathways during days 1-3 after Mtb infection. Confocal microscopic analysis demonstrates that Mtb-infected THP-1 cells or hMDMs rarely exhibited apoptosis. Immunoblotting shows that Mtb induces significant CASP3 and GSDME activation in THP-1 cells, but not in hMDMs. We show that Mtb, in THP-1 cells but not hMDM, induces a significant increase in GSDMD cleavage, a hallmark of pyroptosis. MLKL phosphorylation was not observed in THP-1 cells or hMDMs during Mtb infections, indicating an absence of necroptosis. No changes in ferroptosis markers such as GPX4 expression or lipid peroxidation levels were detected. Time-lapse live-cell imaging revealed no lysosomal membrane permeabilization prior to plasma membrane rupture (PMR). However, we observed DNA release from Mtb-infected THP-1 cells and hMDMs after PMR. The DNA released from THP-1 cells exhibits low levels of myeloperoxidase and histone H3 citrullination. High-resolution confocal imaging shows that Mtb is associated with the released DNA. We demonstrate that pyroptosis induction in THP-1 cells is dispensable for the DNA release and cell death induction. In conclusion, our results reveal that Mtb-triggered cell death in hMDMs bypasses canonical cell death pathways like apoptosis, pyroptosis, necroptosis, and ferroptosis. Instead, cell death in both THP-1 cells and hMDMs correlates with DNA release, potentially similar to NETosis in neutrophils.

DOI: https://doi.org/10.1038/s41420-026-03156-1
EMBO Rep. 2026 Jun 08.

Caspase-8 silences cell death-independent constitutive immune activation driven by tonic TNF-α.

Germana Lentini, Francesco Coppolino, Agata Famà, Giuseppe Valerio De Gaetano, Federica Grasso, Luigi Fiore, Alessia Berbiglia, Silvia Carnevale, Isabella Venza, Egil Lien, Mario Venza, Sebastien Jaillon, Giuseppe Teti, Concetta Beninati.

Caspase-8 is essential for maintaining organismal integrity by preventing cell death and subsequent inflammation in specific epithelial and endothelial tissues. Here, we show that caspase-8 also controls a systemic, cell death-independent inflammatory pathway that is constitutively active during homeostasis. In vivo, selective caspase-8 inhibition produces, in the absence of other stimuli, marked neutrophilia driven by circulating proinflammatory and chemotactic cytokines and promotes bacterial clearance during infection. In vitro, caspase-8 inhibition triggers in neutrophils, but not in macrophages, a profound transcriptional response associated with the release of IL-1β and other cytokines. This process requires tonic TNF-α production by neutrophils, which acts autocrinally to sequentially activate RIPK1, RIPK3, MAPKs, and NF-κB. The IL-1β release induced by caspase-8 inhibition requires gasdermin D and neutrophil serine proteases, but not canonical inflammasome components. Our data uncover the mechanistic features of a neutrophil-centric, proinflammatory pathway that can be therapeutically targeted to augment host defenses against pathogens.

DOI: https://doi.org/10.1038/s44319-026-00813-5
Nat Commun. 2026 Jun 10.

Infection-induced glucose starvation triggers NINJ1-dependent macrophage lysis and Candida escape.

Harshini Weerasinghe, Orawan Tulyaprawat, Helen Stölting, Johannes Sonnberger, Byron Mobbs, Joshua Nickson, Theresa Lange, Bryce van Denderen, Tricia L Lo, Tim Bastian Schille, Françios A B Olivier, Natasha Kapoor-Kaushik, Timothy A Gottschalk, Cathrine Hall, John Silke, James E Vince, Kate E Lawlor, Stefan Bröer, Kate Schroder, Sabrina Sofia Burgener, Bernhard Hube, Thomas Naderer, Adam J Rose, Ana Traven.

Pathogens compete for glucose with macrophages, which disrupts host glycolysis, modulates antimicrobial responses and causes macrophage death. We show that glucose starvation induced by major fungal pathogens Candida albicans and Candida auris causes macrophage lysis by activating NINJ1, the executioner of membrane rupture during cell death. In glucose-starved macrophages, NINJ1 ruptures membranes independently of known cell death programs. Consistently, NINJ1 is the dominant effector of fungal-induced macrophage damage amongst host cell death factors. Supplementation of the amino acid alanine rescues glucose-starved macrophages better than glucose, and it does so by inhibiting NINJ1 oligomerization. Moreover, C. albicans infection disrupts amino acid metabolism in mice and reduces serum alanine. Finally, NINJ1-mediated membrane rupture enables C. albicans egress from macrophages together with the toxin candidalysin. We establish the mechanism of glucose starvation-induced macrophage damage by NINJ1, and demonstrate the roles of NINJ1 and alanine in immune responses to Candida and fungal escape.

DOI: https://doi.org/10.1038/s41467-026-74195-6
Elife. 2026 Jun 10. pii: RP111544. [Epub ahead of print]15

Cell size modulates ferroptosis susceptibility.

Evgeny Zatulovskiy, Magdalena B Murray, Shuyuan Zhang, Scott J Dixon, Jan M Skotheim.

  Size is a fundamental property of cells that influences many aspects of their physiology. This is because cell size sets the scale for all subcellular components and drives changes in the composition of the proteome. Given that large and small cells differ in their biochemical composition, we hypothesized that they should also differ in how they respond to signals and make decisions. Here, we investigated how cell size affects the susceptibility of human cells to cell death. We found that large cells are more resistant to ferroptosis caused by system xc- inhibition. Ferroptosis is a type of cell death characterized by the iron-dependent accumulation of toxic lipid peroxides. This process is opposed by cysteine-dependent lipid peroxide detoxification mechanisms. We found that larger cells exhibit higher concentrations of the cysteine-containing metabolite glutathione and lower concentrations of membrane lipid peroxides. Mechanistically, this can be explained by the fact that larger cells had lower concentrations of an enzyme that enriches cellular membranes with peroxidation-prone polyunsaturated fatty acids, ACSL4, and increased concentrations of the glutathione-producing enzymes glutamate-cysteine ligase and glutathione synthetase, the iron-chelating protein ferritin, and the lysosomal protease cathepsin B, which can catabolize cysteine-rich extracellular proteins to produce additional cystine for fueling the synthesis of glutathione. Taken together, our results highlight the significant impact of cell size on cellular function and survival, revealing a size-dependent vulnerability to ferroptosis that could influence therapeutic strategies based on this cell death pathway.

Keywords:  biochemistry; cell biology; cell death; cell size; chemical biology; erastin2; ferroptosis; glutathione; heterogeneous response; human; scaling

DOI:  https://doi.org/10.7554/eLife.111544
EMBO J. 2026 Jun 09.

GBP1 recruitment to actin-rich pedestals of extracellular Gram-negative bacteria promotes pyroptosis.

Daniel J Bennison, Ishaan Chaudhary, Dharitri Chaudhuri, Justin Chun Ngai Wong, Ayush Punwatkar, Priyanka Biswas, Miyu Stephenson, Qiyun Zhong, Wouter W Kallemeijn, Marianne Guenot, Sandra Koigi, Diana Papp, John P Thomas, Dimple Dixit, Tamas Korcsmaros, Arthur M Talman, Eva-Maria Frickel, Edward W Tate, Sandhya S Visweswariah, Gad Frankel, Avinash R Shenoy.

The IFNγ-induced GTPase guanylate-binding protein 1 (GBP1) binds to lipopolysaccharide (LPS) on cytosolic gram-negative bacteria and promotes pyroptosis via the recruitment and activation of caspase-4 on the bacterial outer membrane. Enteropathogenic and enterohaemorrhagic Escherichia coli (EPEC and EHEC, respectively) are extracellular pathogens that adhere to host cells and stimulate dense actin polymerisation underneath their attachment sites, generating structures described as actin-rich pedestals. Here, we show that GBP1 traffics to actin-rich pedestals in human cells infected with EPEC or EHEC in vitro and mouse colonocytes infected with the EPEC-like murine pathogen Citrobacter rodentium in vivo. GBP1 promotes caspase-4 recruitment to actin-rich pedestals, leading to pyroptosis and IL-18 release. GBP1 mutants defective in LPS coatomer formation also localise to EPEC pedestals. A novel assay that mimics pathogenic effector activity reveals GBP1 recruitment to sterile actin polymerisation sites. We conclude that cytosolic GBP1 is mobilised to sites of pathogen-induced actin remodelling independently of LPS. Our study establishes that GBP1 not only operates as a pattern-recognition receptor but also orchestrates effector-triggered immunity against pathogens that hijack the actin cytoskeleton.

DOI: https://doi.org/10.1038/s44318-026-00830-z
Methods Cell Biol. 2026 ;pii: S0091-679X(26)00079-8. [Epub ahead of print]208 71-83

Time-lapse confocal laser scanning microscopy analysis of FOOD formation.

Ponsuge T M Cooray, Ivan H K Poon, Stephanie F Rutter.

  Apoptotic cells undergo various morphological changes as they die, including membrane blebbing, apoptopodia formation, and the generation of apoptotic bodies. Recently, a novel step in the disassembly of apoptotic adherent cells was described, wherein cells retract from the substrate leaving behind phosphatidylserine and F-actin rich membrane remnants that are tightly anchored to the substrate. Over time, this membrane, coined the 'FOotprint Of Death' or 'FOOD' rounds into extracellular vesicles known as 'FOOD-derived Apoptotic cell derived extracellular vesicles' or 'F-ApoEVs'. Here, we detail an imaging approach to monitor the morphological changes of apoptotic cells as they retract from the extracellular substrate and generate FOOD and F-ApoEVs, using fluorescent conjugated annexin V and actin cytoskeletal stains. With some variations, we expect this approach to be compatible with a range of fluorescent dyes, stains, cell types, and methods of apoptosis induction to detect the formation of FOOD/ F-ApoEV during apoptosis.

Keywords:  Apoptotic cell disassembly; Confocal laser scanning microscopy; Dynamic imaging; F-ApoEVs; The FOotprint Of Death

DOI:  https://doi.org/10.1016/bs.mcb.2026.02.011
RSC Chem Biol. 2026 Jun 04.

Methylation of DNA duplexes regulates cGAS-mediated innate immune activation via condensate formation.

Kunihiko Morihiro, Manami Baba, Moeko Yamada, Akimitsu Okamoto.

Exogenous DNA activates innate immunity via the cGAS-STING pathway, but the impact of chemical modifications has not been investigated sufficiently. Here, we show that DNA duplexes containing 2'-O-methyl ribonucleotides enhance cGAS-driven liquid-liquid phase separation and enzymatic activity, function effectively in DNA nanotherapeutics in living cells, and enable the rational control of innate immune signaling.

DOI: https://doi.org/10.1039/d6cb00059b
Methods Cell Biol. 2026 ;pii: S0091-679X(26)00069-5. [Epub ahead of print]208 221-248

Super-resolution imaging of cell death in Drosophila tissues via expansion and pan-expansion microscopy.

Alina Kolpakova, Keren Yacobi-Sharon, Eli Arama.

  Cell death is essential for the survival and homeostasis of nearly all organisms. Programmed cell death (PCD) originally described the reproducible elimination of specific cells during metazoan development. Apoptosis, the best-studied form of PCD, is characterized by distinct morphological, cellular, and biochemical hallmarks. However, the identification of alternative cell death pathways has highlighted the need to clearly distinguish among them, especially when exploring new paradigms of developmental alternative cell death pathways. Furthermore, the discovery of numerous non-lethal functions of apoptotic caspases, collectively termed non-lethal caspase-dependent processes (CDPs), has underscored the importance of differentiating apoptosis from CDPs within tissues in vivo. Morphological analysis remains essential for differentiating death pathways and for determining whether cells are dying or surviving, yet such analyses traditionally relied on electron microscopy, which is labor-intensive, costly, and technically demanding. Expansion microscopy (ExM), a technique that physically enlarges biological samples isotropically, has transformed light microscopy by enabling super- and ultra-resolution imaging using standard fluorescence or confocal platforms. Here, we describe two ExM-based protocols optimized for, but not limited to, Drosophila tissues: ExM, achieving ∼4× expansion, and pan-ExM, yielding ∼12-16× expansion. ExM supports super-resolution imaging of immunofluorescent labels, whereas pan-ExM provides access to ultrastructural features, albeit with limited capacity for specific protein detection. We also compare apoptotic and non-apoptotic cell death morphologies before and after ExM and pan-ExM. Given the growing diversity of ExM strategies, this chapter offers a comprehensive introduction to these rapidly advancing methodologies.

Keywords:  Apoptosis; Drosophila; Expansion microscopy (ExM); Morphological features of cell death; Non-apoptotic cell death; Pan-expansion microscopy (pan-ExM); Programmed cell death; Super-resolution imaging; Ultrastructural features of cell death

DOI:  https://doi.org/10.1016/bs.mcb.2026.02.004
Methods Cell Biol. 2026 ;pii: S0091-679X(26)00066-X. [Epub ahead of print]208 55-69

Screening and identification of protein-protein interaction using proximity labeling.

Jiang Hua, Francis Ka-Ming Chan.

  Cell death is a fundamental biological process with critical roles in both normal physiology and pathological conditions, especially programmed cell death, such as apoptosis, necroptosis and pyroptosis. Programmed cell death is mediated by cascade signaling transduction rely on protein-protein interaction. Necroptosis mediator RIPK1, RIPK3 and MLKL have been shown to be regulated by different types of post-translational modifications (PTMs), suggesting that additional factors must associate with them during necroptosis. Proximity labeling (PL) has been used to tag and identify proteins, RNAs, or other biomolecules in close proximity (∼10-20 nm) to a target protein of interest. Therefore, utilizing proximity labeling coupled mass spectrometry to identify weak and transient interactors of necroptosis mediators, will be helpful for the further understanding of cell death mechanisms and functions. Here, in this chapter, we provide a step-by-step protocol for using TurboID-based proximity labeling to map interactors and regulators of key necroptotic proteins (RIPK1, RIPK3, MLKL and ZBP1).

Keywords:  Apoptosis; MLKL; Necroptosis; Proximity labeling; RIPK1; RIPK3; ZBP1

DOI:  https://doi.org/10.1016/bs.mcb.2026.02.001
Infect Immun. 2026 Jun 10. e0020026

Staphylococcus aureus inhibits the NLRP3 inflammasome in macrophages to varying degrees during early and late stages of infection.

Saumya Bhagat, Khushpreet Kaur, Jenna R Petronglo, Luke O'Connor, Chun Wang, Yongjia Li, Gaurav Swarnkar, Kunjan Khanna, James E Cassat, Deborah J Veis, Gabriel Mbalaviele.

  Staphylococcus aureus (S. aureus) is a highly virulent pathogen responsible for chronic infections such as osteomyelitis. Although its interaction with the host immune system has been widely studied, the specific role of inflammasomes in regulating the infection within macrophages remains unclear. We investigated this question using bone marrow-derived macrophages (BMDMs) infected with S. aureus for up to 96 hours (h). We observed a significant reduction in intracellular bacterial load beginning at 18 h post-infection (hpi), which continued through 96 hpi; this response was partially attenuated in BMDMs lacking NLRP3. Mechanistically, activation of the NLRP3 inflammasome, including ASC speck formation, IL-1β secretion, and pyroptosis, was modest and detectable only after 18 hpi. Consistent with these findings, infected BMDMs failed to respond robustly to NLRP3 inflammasome activators, lipopolysaccharide (LPS) and nigericin, up to 18 hpi, with only partial restoration of responsiveness at later time points. We also found that NLRP3 deficiency did not significantly affect bacterial burden control or bone homeostasis in experimental S. aureus osteomyelitis. Thus, S. aureus is inadequately controlled by the NLRP3 inflammasome in macrophages and within the bone microenvironment.IMPORTANCEStaphylococcus aureus causes difficult-to-treat, long-lasting infections such as osteomyelitis. While many aspects of how the immune system responds to this pathogen are known, it is still unclear how the inflammasomes contribute to controlling the infection inside macrophages. In this study, we found that macrophages gradually reduced the number of bacteria they contained over time, but this ability was only partly dependent on one major inflammasome component, NLRP3. Importantly, the inflammasome response to S. aureus was weak and delayed, becoming noticeable only well after infection had begun. Even when stimulated with agents that normally trigger a strong inflammasome response, infected macrophages responded poorly. Consistent with these findings, the absence of NLRP3 did not significantly worsen bacterial control or bone damage in a mouse model of osteomyelitis. Together, these results suggest that S. aureus largely evades inflammasome-based defenses, limiting their effectiveness in controlling infection in immune cells and bone.

Keywords:  IL-1β; NLRP3 inflammasome; Staphylococcus aureus; bone marrow-derived macrophages; caspase-1; gasdermin D; host-pathogen interaction; infection; osteomyelitis; pyroptosis

DOI:  https://doi.org/10.1128/iai.00200-26
Proc Natl Acad Sci U S A. 2026 Jun 16. 123(24): e2536989123

Loss of erythrocyte sialic acid in sepsis disrupts inhibitory Siglec interactions, driving neutrophil hyperactivation and NET outspread.

Anna Such, Weronika Ortmann, Gabriela Burczyk, Jacek Czepiel, Monika Bociaga-Jasik, Paweł Link-Lenczowski, Elzbieta Kolaczkowska.

  Systemic inflammation is accompanied by overwhelming neutrophil activation, resulting in extensive formation of neutrophil extracellular traps (NETs). Although NETs are beneficial for pathogen entrapment, they become detrimental; therefore, their endogenous regulation is pivotal for survival. In this context, the observation that healthy individuals' red blood cells (RBCs) can inhibit NET formation induced by lipopolysaccharide (LPS)-primed neutrophils is noteworthy. However, during systemic inflammation, NET formation in the vasculature is robust despite constant neutrophil exposure to RBCs. Herein, we reveal that for RBCs and neutrophils from septic individuals, but not from those with local inflammation, interactions cease and NET inhibition is abolished. This is observed even if only one cell type originates from an inflamed individual, and occurs in both human and murine cells. Moreover, 6 to 10 d post-sepsis induction (mice) or diagnosis (humans), during the resolution of inflammation, interactions are restored and hence fail during the critical stages of inflammation. Mechanistically, this is not due to passive LPS absorption by RBCs, which would limit its availability for neutrophils. In contrast, the cessation of NET inhibition is due to inflammation-induced RBC desialylation (removal of α2,3-linked sialic acids) and decreased expression of inhibitory Siglec-E/9 and -G, but not Siglec-F/5, on neutrophils. Accordingly, concurrent addition of exogenous polysialic acid (PolySia) and pharmacologically controlled accumulation of Siglec molecules on neutrophils preserves the ability of erythrocytes to inhibit PAD4-dependent NET formation upon interaction with neutrophils of endotoxemic mice. These findings open avenues for clinically controlling NET formation during systemic inflammation, given the abundance of erythrocytes in the blood.

Keywords:  erythrocytes; neutrophil extracellular traps (NETs); neutrophils; sepsis; siglec receptors

DOI:  https://doi.org/10.1073/pnas.2536989123
Cells. 2026 May 28. pii: 994. [Epub ahead of print]15(11):

Role of the Host Membrane Trafficking Protein Dynamin 2 in Cell-to-Cell Spread of Bacterial Pathogens.

Keith Ireton.

  Although evolutionarily distant, the bacteria Listeria monocytogenes, Shigella flexneri, and Burkholderia thailandensis each undergo a "cell-to-cell" spreading process that allows these pathogens to disseminate within human tissues. Spread initiates when bacteria polymerize actin filaments that propel them through the host cell cytosol. The pathogens then remodel the plasma membrane into protrusions that are internalized by adjacent cells and resolved into double membranous vacuoles (DMVs) which lyse to liberate bacteria. In this review, we discuss recent publications indicating that L. monocytogenes, S. flexneri, and B. thailandensis each enhance their spread by altering the subcellular localization of human Dynamin 2-a GTPase that regulates endocytosis and other trafficking pathways. Interestingly, Dynamin 2 controls distinct steps in spread of L. monocytogenes, S. flexneri, and B. thailandensis. In the case of L. monocytogenes, the GTPase has the potential to restrict protrusion formation by generating tension at tight junctions. However, L. monocytogenes secretes a protein that relieves this restriction of protrusions, allowing efficient spread. During dissemination of S. flexneri and B. thailandensis, Dynamin 2 is co-opted to resolve protrusions into DMVs. B. thailandensis also mobilizes Dynamin 2 to lyse DMVs. These findings highlight diverse ways in which bacteria control Dynamin 2 to augment spread.

Keywords:  Burkholderia thailandensis; Listeria monocytogenes; Shigella flexneri; cell-to-cell spread; dynamin 2 GTPase

DOI:  https://doi.org/10.3390/cells15110994
Commun Biol. 2026 Jun 08. pii: 773. [Epub ahead of print]9(1):

The signaling and regulatory functions of dying cells and cell corpses.

Guolin Zhang, Jinming Liu, Wenhui Li, Boya Wang, Haoxuan Feng, Tongzhou Cui, Taowen Pan, Dapeng Chen, Yongjian Xiong.

Cells die extensively each day under physiological and pathological conditions, yet dying cells and cell corpses are not merely passive endpoints. Increasing evidence shows that they actively shape their microenvironment during death-program execution, signal release or exposure, corpse processing, and responses of neighboring cells and phagocytes. We here summarize six major roles of dying cells and cell corpses: promoting tissue turnover and remodeling, supporting neighboring-cell survival, mediating intercellular communication, providing metabolic substrates and rewiring phagocyte metabolism, facilitating corpse recognition and clearance, and modulating immune responses. Overall, they provide a broader framework for understanding cell death and identifying therapeutic opportunities.

DOI: https://doi.org/10.1038/s42003-026-10428-0
Front Immunol. 2026 ;17 1759365

ANXA2 promotes NLRP3 inflammasome activation and neuronal pyroptosis after intracerebral hemorrhage.

Qingyuan Wu, Yuetao Wen, Yiqing Shen, XiangYu Chen, WenSong Yang, Limin Ma, Peng Xie.

   Background: Intracerebral hemorrhage (ICH) is a severe form of stroke lacking effective pharmacotherapy, in part because upstream regulators initiating secondary brain injury are not well understood. Pyroptosis mediated by activation of the NLRP3 inflammasome is a major contributor to neuronal death after ICH. However, the upstream mechanisms remain to be fully elucidated.
Methods: We performed integrative transcriptomic-proteomic profiling of mouse ICH brain tissues with in vivo functional validation. Annexin A2 (ANXA2), identified as a hub protein, was silenced via genetic knockdown. Neurological function, brain pathology, and pyroptotic signaling were assessed by behavioral tests, histology, Western blotting, immunofluorescence, and co-immunoprecipitation.
Results: Multi-omics and network analyses identified ANXA2 as a prominently upregulated hub protein after ICH. Co-immunoprecipitation demonstrated an association between ANXA2 and NLRP3, while ANXA2 silencing reduced NLRP3 inflammasome activation, decreased GSDMD cleavage and IL-1β/IL-18 secretion and significantly improved neurological function while alleviating brain injury.
Conclusions: This study reveals a previously unrecognized ANXA2-NLRP3-pyroptosis pathway in ICH, revealing a neuronal-immune convergence mechanism in inflammasome regulation. These findings provide new insight into neuronal pyroptosis after ICH and underscore ANXA2 as a predominantly neuronal factor associated with inflammasome activation in hemorrhagic stroke.

Keywords:  AnxA2; NLRP3 inflammasome; intracerebral hemorrhage; neuroinflammation; pyroptosis

DOI:  https://doi.org/10.3389/fimmu.2026.1759365
Methods Cell Biol. 2026 ;pii: S0091-679X(26)00071-3. [Epub ahead of print]208 203-219

Measuring efferocytosis by intraperitoneal clearance assay.

Caitlin L Vella, Dilara C Ozkocak, Amy A Baxter.

  Efferocytosis is a critical process by which phagocytes identify, engulf and clear apoptotic cells, thereby preserving immune tolerance and tissue homeostasis. While in vitro assays have advanced our understanding of the molecular mechanisms underlying apoptotic cell clearance, they lack the complexity of the physiological environment. This chapter presents a detailed in vivo approach for assessing efferocytosis by mouse peritoneal macrophages using the intraperitoneal (IP) clearance assay. By administering fluorescently labelled apoptotic Jurkat T cells into the peritoneal cavity, this method enables the quantification of apoptotic cell uptake in a native tissue context, capturing the influence of local immune signals, stromal interactions and systemic factors. This protocol is compatible with genetic and pharmacological manipulations, allowing researchers to investigate how various interventions affect efferocytic capacity. Overall, this approach provides a robust and physiologically relevant framework for studying efferocytosis and its implications in both homeostatic and pathological settings.

Keywords:  Apoptosis; Efferocytosis; Flow cytometry; Intraperitioneal clearance; Phagocytes

DOI:  https://doi.org/10.1016/bs.mcb.2026.02.006
bioRxiv. 2026 Jun 02. pii: 2026.05.31.728998. [Epub ahead of print]

Cell fusion after wounding requires plasma membrane damage and endocytosis.

Junmin Hua, Evan S Krystofiak, Andrew D Pumford, Andrea Page-McCaw, M Shane Hutson.

Tissue wounds comprise both dead and damaged cells. In epithelial wounds, repair is accomplished by cells at the wound edges, which are themselves often damaged. In the Drosophila pupal notum, wound-adjacent epithelial cells with plasma membrane damage often fuse to form syncytia; when plasma membrane damage is prevented, syncytia do not form. Damaged cells share cytoplasm as soon as milliseconds after wounding, and fusion pores connecting cell membranes form minutes later. A genetic screen reveals that wound-induced fusion requires endocytosis machinery, and dynamin localization indicates that endocytosis preferentially targets plasma membrane removed during fusion. Endocytosis promotes cell fusion by specifically promoting fusion pore expansion, indicated by quantitative analysis of cytoplasmic sharing between cells over time. Without endocytosis-mediated cell fusion, wound healing is slowed. Together, our results support a model of damage-induced cell fusion in which plasma membrane damage initiates fusion pores and endocytosis expands fusion pores, resulting in cellular fusion as an integration of single cell damage with tissue repair.

DOI: https://doi.org/10.64898/2026.05.31.728998
Autophagy. 2026 Jun 12. 1-3

Cell stress and death liberate the autophagy-inhibitory tissue stress hormone DBI/ACBP into the circulation.

Yan Rong, Flavia Lambertucci, Yaning Yang, Yanbing Dong, Maria Chiara Maiuri, Isabelle Martins, Guido Kroemer.

  Autophagy constitutes a major adaptive response that preserves cellular and organismal homeostasis during stress. However, stress responses also engage systemic communication pathways that may either maintain resilience or propagate pathology. We previously identified acyl-CoA-binding protein, also known as diazepam-binding inhibitor (DBI/ACBP), as a phylogenetically conserved extracellular factor secreted by stressed cells through an unconventional autophagy-dependent pathway. Once released, extracellular DBI/ACBP acts as a feedback inhibitor of autophagy and promotes metabolic and inflammatory alterations. In our most recent work, we identify regulated cell death as an additional major mechanism responsible for extracellular DBI/ACBP accumulation. Plasma DBI/ACBP concentrations correlate with markers of inflammation, senescence and multiorgan dysfunction in hospitalized patients. Experimentally induced injury to liver, kidney, pancreas or skeletal muscle indistinguishably causes rapid increases in circulating DBI/ACBP. Mechanistically, apoptosis, ferroptosis and necroptosis all provoke loss of intracellular DBI/ACBP together with its extracellular release following plasma membrane permeabilization. Pharmacological inhibition of these death pathways suppresses DBI/ACBP liberation. Across large human cohorts, elevated plasma DBI/ACBP is associated with aging, systemic inflammation, multiorgan dysfunction and future morbidity. We propose that DBI/ACBP is not merely a biomarker of tissue damage but rather a systemic autophagy-inhibitory stress signal contributing to maladaptive interorgan communication during aging and disease.

Keywords:  Aging; disease; mortality; organ failure; stress

DOI:  https://doi.org/10.1080/15548627.2026.2685761
Int J Mol Sci. 2026 Jun 04. pii: 5075. [Epub ahead of print]27(11):

Toll-like Receptor 3 as a Context-Dependent Molecular Switch in Epithelial Cancers: Balancing Cell Death and Tumor-Supportive Programs.

Amarilis Pérez-Baños, Andrés Tittarelli.

  Toll-like receptor 3 (TLR3) is a double-stranded RNA sensor that plays a dual and context-dependent role in epithelial cancers, promoting either regulated cell death (RCD) or tumor-supportive programs. While TLR3 activation has been widely explored for its capacity to induce apoptosis or necroptosis and enhance antitumor immunity, accumulating evidence indicates that cancer cells can use TLR3 signaling to sustain proliferation, migration, stemness, and therapy resistance. In this review, we provide a comprehensive and mechanistic analysis of TLR3 signaling in epithelial cancers, encompassing canonical and non-canonical modules that regulate cancer cell fate. We examine how TLR3 activation engages interconnected RCD programs, including apoptosis, necroptosis, and immunogenic cell death, and contrast these with pathways driving tumor plasticity and progression. Importantly, we discuss the key determinants governing TLR3 signaling output, including signaling complex composition; ligand origin and delivery; TLR3 subcellular localization; and cancer cell-intrinsic factors such as genetic, epigenetic, and metabolic states. We propose an integrative framework in which TLR3 signaling influences cancer cell fate according to cellular and microenvironmental context.

Keywords:  cancer; regulated cell death; signaling pathways; toll-like receptor 3; tumor cell plasticity

DOI:  https://doi.org/10.3390/ijms27115075
J Immunol. 2026 06 07. pii: vkag122. [Epub ahead of print]215(6):

IFITM1 differentially regulates antibacterial immunity and immunopathology but is dispensable for antiparasitic responses.

Sibongiseni K L Poswayo, Mumin Ozturk, Rudranil Hazra, Antony Fearns, Christophe J Queval, Narjis Thawer, Julius E Chia, Agnes Malobela, Reto Guler, Ramona Hurdayal, Frank Brombacher, Maximiliano G Gutierrez, Suraj P Parihar.

  Interferon-induced transmembrane (IFITM) proteins underpin antiviral responses, yet their role in bacterial infections remains underexplored, particularly for parasites. We probed the role of IFITM1 in Mycobacterium tuberculosis (Mtb), Listeria monocytogenes (Lm), and Leishmania major infection using IFITM1 knockout mice. Notably, IFITM1 was upregulated in murine and human macrophages, as well as in PBMCs during active tuberculosis (TB), subsiding with therapy. IFITM1 also accumulated in the lungs of outbred mice and macaques that progressed to TB. IFITM1-deficient mice displayed no differences in the acute phase; however, chronic Mtb infection revealed lower bacterial loads, mitigated lung pathology, dampened inflammatory cell recruitment, and decreased cytokines. IFITM1-/- macrophages curbed intracellular H37RV and HN878 growth, skewing proinflammatory cytokine (IL-1α, IL-1β, IL-6, and nitric oxide) production while phagosome maturation and autophagy remained unaffected. Furthermore, HN878-infected IFITM1-/- mice exhibited increased lung cell death by TUNEL staining, driving enhanced mortality. Lm similarly increased IFITM1 expression in macrophages, liver, and spleen. IFITM1-/- mice exhibited reduced early tissue burdens and serum IFN-γ, TNF, and IL-6, yet liver pathology escalated, driving mortality and suggesting dysregulated inflammation. These macrophages also limited intracellular Lm growth, with increased necrosis. By contrast, L. major footpad swelling and parasitic loads remained unaffected in knockout animals. Together, IFITM1 exacerbates TB and listeriosis pathology by calibrating inflammation against bacterial control, but plays no role in cutaneous leishmaniasis. These findings reveal IFITM1-specific contributions to bacterial but not parasitic infections, favoring disease tolerance.

Keywords:  IFITM1; knockout mice; leishmaniasis; listeriosis; tuberculosis

DOI:  https://doi.org/10.1093/jimmun/vkag122
J Cell Biol. 2026 Aug 03. pii: e202603170. [Epub ahead of print]225(8):

Met-Vision reveals coexisting energetic states in tissue macrophages redistributed by inflammation.

Adriana Lecourieux, Margot Bardou, Zacarias Garcia, Philippe Bousso.

The function of tissue-associated macrophages is tightly linked to their energy metabolism. Yet, the diversity of macrophage metabolic profiles coexisting in tissues at homeostasis or during immune challenges is incompletely understood. Here, we introduce Met-Vision, an imaging-based pipeline for single-cell functional profiling and classification of energy metabolism. Across multiple tissue contexts, we identified that macrophages do not adopt a uniform metabolic profile but typically coexist in four discrete metabolic states with distinct dependence on OXPHOS and metabolic plasticity. Inflammation reconfigured the distribution of macrophage metabolic profiles that remained heterogeneous. Notably, inflammation-derived nitric oxide finely tuned the distribution of macrophage energetic states. These findings challenge the view of homogeneous metabolic activation and reveal a layer of metabolic diversity in tissue at steady state and during inflammation. The ability to stratify macrophage energy metabolic profiles with Met-Vision should help guide the development of metabolism-targeted therapies for inflammatory diseases, cancer, and metabolic disorders.

DOI: https://doi.org/10.1083/jcb.202603170
Cell Death Discov. 2026 Jun 10.

Mechanistic analysis of MLKL-driven cell survival.

Peijia Jiang, Xiaoyang Liu, Mauricio J Reginato, Kirill V Rosen.

MLKL pseudokinase is a critical executioner of necroptotic cell death. MLKL drives necroptosis by forming pores in the cell membrane. A growing body of data indicates that, in addition to this well-established role, MLKL can promote cell survival in certain contexts. Moreover, pharmacological or genetic MLKL inhibition was shown to suppress in vivo growth of several tumor types. It was found that MLKL protects cancer cells from various cell death-inducing stimuli by promoting autophagy or preserving the mitochondrial function of the cells. It was proposed that both of these MLKL effects prevent parthanatos, a cell death type mediated by hyperactivation of PARP1 and subsequent PARP1-dependent chromosomal DNA degradation. In addition, MLKL was found to protect tumor cells from the death receptor-induced demise and trigger the secretion of the growth-promoting cytokines by the cells. Notably, MLKL-deficient mice are healthy, while pharmacological MLKL inhibitors are not significantly toxic to mice. Hence, targeting MLKL in vivo to block MLKL-dependent cancer cell survival is feasible. The mechanisms of the pro-survival MLKL effects is the subject of this review.

DOI: https://doi.org/10.1038/s41420-026-03187-8
J Invest Dermatol. 2026 Jun 09. pii: S0022-202X(26)01050-X. [Epub ahead of print]

The role of neutrophils and neutrophil extracellular traps in Staphylococcus aureus colonization in healthy and inflamed skin.

Jule Riebelmann, Birgit Schittek.

  Staphylococcus aureus (S aureus) commonly causes skin infections and is abundant on the skin of patients with atopic dermatitis, where it worsens inflammation and drives skin barrier defects. Neutrophils help to control S aureus infection through their antimicrobial activity and by recruiting other immune cells; however, they can also promote S aureus skin colonization. Excessive neutrophil activity and release of neutrophil extracellular traps may impair the skin barrier and thereby promote colonization. Moreover, S aureus has evolved strategies to evade neutrophil defenses. This review explores neutrophil-skin interactions in healthy and inflamed skin and potential therapeutic strategies targeting these interactions to reduce S aureus colonization in diseases like atopic dermatitis.

Keywords:  Atopic dermatitis; Inflammatory skin diseases; Neutrophil extracellular traps; Neutrophils; Staphylococcus aureus

DOI:  https://doi.org/10.1016/j.jid.2026.04.006
Front Mol Biosci. 2026 ;13 1732916

Mechanisms driving open-closed transitions and dimer stability of the large GTPases hGBP1 and hGBP5.

Bastian F Bundschuh, Jennifer Loschwitz, Birgit Strodel.

  The human guanylate-binding proteins 1 and 5 (hGBP1/5) are key players in innate immunity, vital for defending against intracellular pathogens and mediating membrane-associated immune responses. These protein functions are closely linked to their structural dynamics, making a detailed understanding of the conformational behavior imperative. The closed conformation of hGBP1 is well-characterized through crystallography, but the structural basis for its transition to an active, open state remains less understood. This study uses all-atom and coarse-grained molecular dynamics simulations to investigate the stability and motions of extended monomeric and dimeric hGBP1 and hGBP5 with and without GTP bound. Results reveal that dimers exhibit greater stability than monomers, primarily due to extensive stalk interactions that facilitate a structural crossing of the protomers at the interface of the GTPase and middle domains. This arrangement aligns the middle and effector domains parallel to one another, further stabilizing the dimeric state through the formation of coiled-coil structures supported by salt bridges and hydrophobic contacts. Notably, monomers of both hGBP1 and hGBP5 can revert to a closed state stabilized by a network of salt bridges between the effector domain and the surrounding domains. In hGBP5, this transition is further facilitated by the geranylgeranyl group, which more effectively reaches and buries itself within a hydrophobic pocket of the GTPase domain compared to hGBP1. These findings highlight key factors affecting the stability of hGBP1/5 monomers and dimers, providing insights into their activation mechanisms that are relevant for their role in innate immunity.

Keywords:  coiled-coil formation; dynamin-like proteins (DLPs); guanylate-binding proteins (GBPs); large GTPases; molecular dynamics simulations (MD); open-closed transition; protein dimerization

DOI:  https://doi.org/10.3389/fmolb.2026.1732916
Biochem Soc Trans. 2026 Jun 24. 54(6): 745-754

Ubiquitin beyond the proteome: lipids, glycans, metabolites, nucleic acids, and an expanding molecular landscape.

Akinori Endo, Yukiko Yoshida.

  Ubiquitination is a versatile post-translational modification process in which the small globular protein ubiquitin is covalently attached to substrate proteins to generate diverse cellular signals. Although originally characterized by its role in proteasome-mediated protein degradation, ubiquitination is now recognized as a central regulator of numerous processes, including signaling, trafficking, and immunity. Canonical ubiquitination is mediated by a cascade of E1 (activating), E2 (conjugating), and E3 (ligase) enzymes that repeatedly conjugate ubiquitin molecules to lysine residues on substrate proteins, leading to the formation of polyubiquitin chains with distinct topologies. The modification is reversed by deubiquitinating enzymes. Notably, components of the ubiquitin system comprise approximately 7% of the human proteome, underscoring its importance in biological regulation. Recent advances have revealed the broad scope of ubiquitination. Ubiquitin was found to conjugate not only to lysine but also to serine, threonine, and cysteine, indicating its unexpected chemical flexibility. Furthermore, ubiquitination can be directed toward other post-translational modifications, particularly glycosylation and ADP-ribosylation, highlighting the extensive crosstalk between modification systems. Strikingly, lipids, sugars, metabolites, nucleic acids, and even synthetic small-molecule compounds have been identified as ubiquitinated substrates. The hypothesis that virtually all classes of molecules are targeted by ubiquitination has become increasingly plausible. Taken together, these findings redefine ubiquitination as a far more general modification process than previously appreciated. In this mini-review, we focus on recent progress in non-proteinaceous ubiquitination research, summarize emerging substrate classes, and discuss key challenges in elucidating the underlying mechanisms and physiological roles of this expanding modification landscape.

Keywords:  Ubiquitin; Ubiquitin ligases; non-proteinaceous

DOI:  https://doi.org/10.1042/BST20260081
Autophagy. 2026 Jun 13.

STING1 senses mitochondrial damage to promote mitophagy.

Ze-Bo Huang, Jin-Yi Lin, Ling-Jun Cheng, Hayden Weng Siong Tan, Han-Ming Shen, Guang Lu.

  The cGAS-STING1 pathway is essential for innate immunity, while its functions beyond immune activation have emerged as a key research topic. Recent studies have revealed the non-canonical roles of this pathway in autophagy. However, whether it participates in organelle quality control through selective autophagy processes such as mitophagy remains largely unexplored. In our study, we identify the cGAS-STING1 pathway as an essential upstream regulator of PINK1-PRKN-dependent mitophagy. We demonstrate that upon mitochondrial damage, STING1 is recruited to damaged mitochondria in a process requiring PINK1- and VCP/p97-mediated degradation of outer mitochondrial membrane proteins. STING1 at damaged mitochondria then activates TBK1, which phosphorylates the mitophagy receptor OPTN at Ser177, enhancing its recruitment to damaged mitochondria and driving efficient mitophagy. Disruption of the STING1-TBK1-OPTN axis impairs mitophagy and shifts the cellular response from pro-survival mitophagy to apoptosis. Our findings therefore uncover a non-canonical, pro-survival function of the cGAS-STING1 pathway in mitophagy, extending its role beyond innate immunity to the regulation of selective autophagy and cell fate decisions. Abbreviations: BafA1: bafilomycin A1; cGAS: cyclic GMP‑AMP synthase; ER: endoplasmic reticulum; GABARAP: GABA type A receptor-associated protein; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MQC: mitochondrial quality control; mtDNA: mitochondrial DNA; NAC: N-Acetylcysteine; Nec-1: Necrostatin-1; OMM: outer mitochondrial membrane; OPTN: optineurin; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; RIPK1: receptor interacting serine/threonine kinase 1; ROS: reactive oxygen species; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK binding kinase 1; TFEB: transcription factor EB; VCP/p97: valosin containing protein; Z-VAD-FMK: benzyloxycarbony (Cbz)-l-ValAla-Asp (OMe)-fluoromethylketone.

Keywords:  Cell death; OPTN; PINK1-PRKN-dependent mitophagy; cGAS-STING1 pathway; innate immunity; mitochondrial quality control

DOI:  https://doi.org/10.1080/15548627.2026.2689463