Structural mechanisms for regulation of GSDMB pore-forming activity

Zhong, Xiaojing; Zeng, Huan; Zhou, Zhiwei; Su, Ya; Cheng, Hang; Hou, Yanjie; She, Yang; Feng, Na; Wang, Jia; Shao, Feng; Ding, Jingjin

doi:10.1038/s41586-023-05872-5

Cited by 65 publications

(67 citation statements)

References 39 publications

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“…Unexpectedly, oligomers were detected in all noncytotoxic GSDMB-NTs, including B1, B2, B3S, and B5, suggesting that the belt does not markedly contribute to GSDMB-NT oligomerization. Our results contrast with mutagenesis experiments and molecular dynamics simulations pointing toward the involvement of the belt in GSDMB-NT oligomerization (30,31,33).…”

Section: Noncytotoxic Gsdmb-nts Are Incapable Of Membrane Insertioncontrasting

confidence: 99%

“…It is possible that there are subtle lipid binding differences under the detection limit of our assay, because triple charge-reversal mutations of R225-K227-K229 modestly lowered the activity of GSDMB3 in liposomebased experiments (30). Nonetheless, less-aggressive double mutations of R225-K227 to alanines did not affect GSDMB3 activity (31), supporting our postulation that the belt promotes pore formation mainly by an alternative mechanism. Besides the lipid strip results, all GSDMB-NTs contain the hydrophobic anchor and three positively charged patches previously identified as crucial for membrane binding by GSDM-NTs (9,26,32).…”

Section: Noncytotoxic Gsdmb-nts Are Incapable Of Membrane Insertionsupporting

confidence: 76%

“…Therefore, the belt is not part of the disordered linker between the NT and CT domains of GSDMs but rather is an ordered, integral structure within the active NT domain. Cryo-EM structures of the membrane-inserted GSDMB (30,31) published during the revision of this manuscript confirmed the orderedness of the belt.…”

Section: A Belt Motif In Gsdm-nt Promotes Pore Formationsupporting

confidence: 65%

“…2D). Consistently, crystal structures of GSDMB5 showed that the belt is disordered in this isoform (30)(31)(32). Therefore, the belt motif in isoforms 3L and 4, which resembles the stabilizing structures formed in GSDMD and GSDMA3, is necessary for pore formation and unstable in the noncytotoxic isoforms.…”

Section: A Belt Motif In Gsdm-nt Promotes Pore Formationsupporting

confidence: 52%

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Alternative splicing of GSDMB modulates killer lymphocyte–triggered pyroptosis

et al. 2023

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Granzyme A from killer lymphocytes cleaves gasdermin B (GSDMB) and triggers pyroptosis in targeted human tumor cells, eliciting antitumor immunity. However, GSDMB has a controversial role in pyroptosis and has been linked to both anti- and protumor functions. Here, we found that GSDMB splicing variants are functionally distinct. Cleaved N-terminal (NT) fragments of GSDMB isoforms 3 and 4 caused pyroptosis, but isoforms 1, 2, and 5 did not. The nonfunctional isoforms have a deleted or modified exon 6 and therefore lack a stable belt motif. The belt likely contributes to the insertion of oligomeric GSDMB-NTs into the membrane. Consistently, noncytotoxic GSDMB-NTs blocked pyroptosis caused by cytotoxic GSDMB-NTs in a dominant-negative manner. Upon natural killer (NK) cell attack, GSDMB3-expressing cells died by pyroptosis, whereas GSDMB4-expressing cells died by mixed pyroptosis and apoptosis, and GSDMB1/2-expressing cells died only by apoptosis. GSDMB4 partially resisted NK cell-triggered cleavage, suggesting that only GSDMB3 is fully functional. GSDMB1-3 were the most abundant isoforms in the tested tumor cell lines and were similarly induced by interferon-γ and the chemotherapy drug methotrexate. Expression of cytotoxic GSDMB3/4 isoforms, but not GSDMB1/2 isoforms that are frequently up-regulated in tumors, was associated with better outcomes in bladder and cervical cancers, suggesting that GSDMB3/4-mediated pyroptosis was protective in those tumors. Our study indicates that tumors may block and evade killer cell-triggered pyroptosis by generating noncytotoxic GSDMB isoforms. Therefore, therapeutics that favor the production of cytotoxic GSDMB isoforms by alternative splicing may improve antitumor immunity.

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Section: Noncytotoxic Gsdmb-nts Are Incapable Of Membrane Insertioncontrasting

confidence: 99%

Section: Noncytotoxic Gsdmb-nts Are Incapable Of Membrane Insertionsupporting

confidence: 76%

Section: A Belt Motif In Gsdm-nt Promotes Pore Formationsupporting

confidence: 65%

Section: A Belt Motif In Gsdm-nt Promotes Pore Formationsupporting

confidence: 52%

See 2 more Smart Citations

Alternative splicing of GSDMB modulates killer lymphocyte–triggered pyroptosis

et al. 2023

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“…Interestingly, VHHGSDMD-2, but not VHHGSDMD-6, binds to a similar surface patch on the GSDMD NTD as the Shigella flexneri ubiquitin ligase IpaH7.8, suggesting mutually exclusive binding modes. IpaH7.8, which targets GSDMD for degradation to prevent pyroptosis and enable infection 44 , has recently been shown to bind to the NTD of both GSDMB and GSDMD through its LRR domain 45,46 . However, as the nanobody binds to GSDMD with a significantly higher affinity than IpaH7.8, displacement of the ubiquitin ligase should prevent its degradation, underlining its application as a biological tool, e.g.…”

Section: Discussionmentioning

confidence: 99%

Pyroptosis inhibiting nanobodies block Gasdermin D pore formation

Kopp

Hagelueken

Jamitzky

et al. 2023

Preprint

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Gasdermin D (GSDMD) is a key mediator of pyroptosis, a pro-inflammatory form of cell death occurring downstream of inflammasome activation as part of the innate immune defence. Upon cleavage by inflammatory caspases, the N-terminal domain of GSDMD forms pores in the plasma membrane resulting in cytokine release and eventually cell death. Targeting GSDMD is an attractive way to dampen inflammation. In this study, six GSDMD targeting nanobodies were characterized in terms of their binding affinity, stability, and effect on GSDMD pore formation. Three of the nanobodies inhibited GSDMD pore formation in a liposome leakage assay, although caspase cleavage was not perturbed. We determined the crystal structure of human GSDMD in complex with two nanobodies at 1.9 Ang resolution, providing detailed insights into the GSDMD-nanobody interactions and epitope binding. The pore formation is sterically blocked by one of the nanobodies that binds to the oligomerization interface of the N-terminal domain in the multi-subunit assembly. Our biochemical and structural findings provide new tools for studying inflammasome biology and build a framework for the design of novel GSDMD targeting drugs.

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Biomaterials Elicit Pyroptosis Enhancing Cancer Immunotherapy

Zhang,

Wang,

Han

et al. 2023

Adv Funct Materials

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Cancer immunotherapy has the potential to revolutionize the treatment of malignant tumors, but its effectiveness is limited by the low immune response rate and immune‐related adverse events. Pyroptosis, as an inflammatory programmed cell death type, triggers strong acute inflammatory response and antitumor immunity, converting “cold” tumors to “hot”. Particularly, biomaterials loading pyroptosis inducers targeting the tumor microenvironment to engineer pyroptosis, have achieved great progress in recent years. Herein, the design strategy, mechanism pathway, and role of biomaterials to induce pyroptosis in cancer immunotherapy are comprehensively reviewed. The present review focuses on the application of biomaterials‐induced pyroptosis in cancer immunotherapy, including nanogel, polymer prodrug, nanovesicle, and mesoporous material. Additionally, the synthesis of a series of stimuli‐responsive nanoplatforms, including glutathione‐responsive, pH‐responsive, reactive oxygen species‐responsive, and enzyme‐mimicking catalytic performance, is described. Meanwhile, it augments multiple immune response processes of cell uptake, antigen presentation, T‐cell activation, and expansion. Finally, the perspectives of pyroptosis‐mediated inflammation to break through the tumor vascular basement membrane barrier achieving efficient volcanic penetration of biomaterials are discussed. Artificial intelligence, multi‐omics analysis, and anthropogenic animal models of organoids are presented, aiming to provide guidance and assistance for constructing effective and controllable pyroptosis‐engineered biomaterials and improving tumor immunotherapy.

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Structural mechanisms for regulation of GSDMB pore-forming activity

Cited by 65 publications

References 39 publications

Alternative splicing of GSDMB modulates killer lymphocyte–triggered pyroptosis

Alternative splicing of GSDMB modulates killer lymphocyte–triggered pyroptosis

Pyroptosis inhibiting nanobodies block Gasdermin D pore formation

Biomaterials Elicit Pyroptosis Enhancing Cancer Immunotherapy

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