Protein engineering reveals that gasdermin A preferentially targets mitochondrial membranes over the plasma membrane during pyroptosis

Kondolf, Hannah; D'Orlando, Dana A; Dubyak, George R.; Abbott, Derek W.

doi:10.1016/j.jbc.2023.102908

Cited by 20 publications

(20 citation statements)

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“…In this sense, GSDMB NT dual impact on membrane and mitochondria mimics the effect of other GSDMs NTs (GSDMA/A3/D/E) [ 14 , 43 , 60 , 65 ]. Recent data indicate that GSDM mitochondrial impairment generally precedes plasma membrane targeting [ 14 , 65 ], but then cell membrane pores are sufficient to cause cellular lysis [ 66 ]. Depending on the GSDM member and biological context, early mitochondrial damage can activate other cooperative cell death mechanisms, like apoptosis (GSDMA and GSDME; [ 12 , 66 ]), autophagic cell death (GSDMA3; [ 43 ]), or necroptosis (GSDMD; [ 67 ]).…”

Section: Discussionmentioning

confidence: 99%

“…Recent data indicate that GSDM mitochondrial impairment generally precedes plasma membrane targeting [ 14 , 65 ], but then cell membrane pores are sufficient to cause cellular lysis [ 66 ]. Depending on the GSDM member and biological context, early mitochondrial damage can activate other cooperative cell death mechanisms, like apoptosis (GSDMA and GSDME; [ 12 , 66 ]), autophagic cell death (GSDMA3; [ 43 ]), or necroptosis (GSDMD; [ 67 ]). We showed that, like GSDMA and GSMDA3 NTs [ 43 , 66 ], GSDMB NT mostly accumulated in mitochondria, but this did not lead to secondary apoptosis (caspase3/7 activation) or increased mitophagy (data not shown).…”

Section: Discussionmentioning

confidence: 99%

“…Depending on the GSDM member and biological context, early mitochondrial damage can activate other cooperative cell death mechanisms, like apoptosis (GSDMA and GSDME; [ 12 , 66 ]), autophagic cell death (GSDMA3; [ 43 ]), or necroptosis (GSDMD; [ 67 ]). We showed that, like GSDMA and GSMDA3 NTs [ 43 , 66 ], GSDMB NT mostly accumulated in mitochondria, but this did not lead to secondary apoptosis (caspase3/7 activation) or increased mitophagy (data not shown). Besides, HEK293 cells are necroptosis-deficient [ 68 ] and lack other endogenous GSDMs [ 17 ], thus GSDMB NT pyroptosis might to be self-sufficient (does not require other secondary cell death mechanisms) and involves the concomitant effect of mitochondrial damage and cell lysis.…”

Section: Discussionmentioning

confidence: 99%

“…Besides, HEK293 cells are necroptosis-deficient [ 68 ] and lack other endogenous GSDMs [ 17 ], thus GSDMB NT pyroptosis might to be self-sufficient (does not require other secondary cell death mechanisms) and involves the concomitant effect of mitochondrial damage and cell lysis. Nonetheless, since GSDM-mediated mitochondrial impairment and cell rupture proceed very quick [ 66 ] to uncover the precise intracellular kinetics and functional consequences of GSDMB activation further studies are required using additional approaches in which GSDMB NT release could be controlled in a more accurate way.…”

Section: Discussionmentioning

confidence: 99%

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Distinct GSDMB protein isoforms and protease cleavage processes differentially control pyroptotic cell death and mitochondrial damage in cancer cells

et al. 2023

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Gasdermin (GSDM)-mediated pyroptosis is functionally involved in multiple diseases, but Gasdermin-B (GSDMB) exhibit cell death-dependent and independent activities in several pathologies including cancer. When the GSDMB pore-forming N-terminal domain is released by Granzyme-A cleavage, it provokes cancer cell death, but uncleaved GSDMB promotes multiple pro-tumoral effects (invasion, metastasis, and drug resistance). To uncover the mechanisms of GSDMB pyroptosis, here we determined the GSDMB regions essential for cell death and described for the first time a differential role of the four translated GSDMB isoforms (GSDMB1-4, that differ in the alternative usage of exons 6-7) in this process. Accordingly, we here prove that exon 6 translation is essential for GSDMB mediated pyroptosis, and therefore, GSDMB isoforms lacking this exon (GSDMB1-2) cannot provoke cancer cell death. Consistently, in breast carcinomas the expression of GSDMB2, and not exon 6-containing variants (GSDMB3-4), associates with unfavourable clinical-pathological parameters. Mechanistically, we show that GSDMB N-terminal constructs containing exon-6 provoke cell membrane lysis and a concomitant mitochondrial damage. Moreover, we have identified specific residues within exon 6 and other regions of the N-terminal domain that are important for GSDMB-triggered cell death as well as for mitochondrial impairment. Additionally, we demonstrated that GSDMB cleavage by specific proteases (Granzyme-A, Neutrophil Elastase and caspases) have different effects on pyroptosis regulation. Thus, immunocyte-derived Granzyme-A can cleave all GSDMB isoforms, but in only those containing exon 6, this processing results in pyroptosis induction. By contrast, the cleavage of GSDMB isoforms by Neutrophil Elastase or caspases produces short N-terminal fragments with no cytotoxic activity, thus suggesting that these proteases act as inhibitory mechanisms of pyroptosis. Summarizing, our results have important implications for understanding the complex roles of GSDMB isoforms in cancer or other pathologies and for the future design of GSDMB-targeted therapies.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Distinct GSDMB protein isoforms and protease cleavage processes differentially control pyroptotic cell death and mitochondrial damage in cancer cells

et al. 2023

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“…The N-terminal functional domain can regulate mitochondrial homeostasis, specifically via mitochondrial oxidative stress and Hsp90/Trap1/Tom70 axis, culminating in mitochondrial permeability transition pore (mPTP) opening and mitochondrial collapse. This response is alleviated by the presence of the autoinhibitory GSDMA C-terminal domain, or pharmacological intervention at the points of ROS, mPTP activation or mitochondrial translocation, and an independent study showed GSDMA has a preference for mitochondrial membranes over plasma membranes ( 95 ). Using overexpressed N-terminal GSDMD and GSDMA, there was a large differential between the two with regards to pyroptotic response, measured by propidium iodide and lactate dehydrogenase release assays.…”

Section: Gsdmamentioning

confidence: 99%

Gasdermins assemble; recent developments in bacteriology and pharmacology

et al. 2023

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The discovery of gasdermin D (GSDMD) as the terminal executioner of pyroptosis provided a large piece of the cell death puzzle, whilst simultaneously and firmly putting the gasdermin family into the limelight. In its purest form, GSDMD provides a connection between the innate alarm systems to an explosive, inflammatory form of cell death to jolt the local environment into immunological action. However, the gasdermin field has moved rapidly and significantly since the original seminal work and novel functions and mechanisms have been recently uncovered, particularly in response to infection. Gasdermins regulate and are regulated by mechanisms such as autophagy, metabolism and NETosis in fighting pathogen and protecting host. Importantly, activators and interactors of the other gasdermins, not just GSDMD, have been recently elucidated and have opened new avenues for gasdermin-based discovery. Key to this is the development of potent and specific tool molecules, so far a challenge for the field. Here we will cover some of these recently discovered areas in relation to bacterial infection before providing an overview of the pharmacological landscape and the challenges associated with targeting gasdermins.

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Non‐viral Gene Therapy for Melanoma Using Lysenin from Eisenia Foetida

Ren,

Yang,

et al. 2024

Advanced Science

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Earthworms, long utilized in traditional medicine, serve as a source of inspiration for modern therapeutics. Lysenin, a defensive factor in the coelom fluid of the earthworm Eisenia fetida, has multiple bioactivities. However, the inherent toxicity of Lysenin as a pore‐forming protein (PFP) restricts its application in therapy. Here, a gene therapy strategy based on Lysenin for cancer treatment is presented. The formulation consists of polymeric nanoparticles complexed with the plasmid encoding Lysenin. After transfection in vitro, melanoma cells can express Lysenin, resulting in necrosis, autophagy, and immunogenic cell death. The secretory signal peptide alters the intracellular distribution of the expressed product of Lysenin, thereby potentiating its anticancer efficacy. The intratumor injection of Lysenin gene formulation can efficiently kill the transfected melanoma cells and activate the antitumor immune response. Notably, no obvious systemic toxicity is observed during the treatment. Non‐viral gene therapy based on Lysenin derived from Eisenia foetida exhibits potential in cancer therapy, which can inspire future cancer therapeutics.

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Protein engineering reveals that gasdermin A preferentially targets mitochondrial membranes over the plasma membrane during pyroptosis

Cited by 20 publications

References 50 publications

Distinct GSDMB protein isoforms and protease cleavage processes differentially control pyroptotic cell death and mitochondrial damage in cancer cells

Distinct GSDMB protein isoforms and protease cleavage processes differentially control pyroptotic cell death and mitochondrial damage in cancer cells

Gasdermins assemble; recent developments in bacteriology and pharmacology

Non‐viral Gene Therapy for Melanoma Using Lysenin from Eisenia Foetida

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