Biology subjects

Ball, D. P.

Publications and source records attributed to Ball, D. P..

7 recordsLinked to original sources

bioRxiv2026

Oxidized TRX1 and DPP8/9 stabilize the MIA40-AIFM1 complex during intrinsic apoptosis

Mammalian cells execute diverse death programs with distinct immunological consequences. Mitochondrial outer membrane permeabilization (MOMP) drives typically immunologically silent intrinsic apoptosis by releasing cytochrome c and other proapoptotic proteins into the cytosol. Here, we show that MOMP also engages two established repressors of the pyroptosis-inducing NLRP1 inflammasome, oxidized thioredoxin-1 (oxTRX1) and catalytically active DPP8/9, in a coordinated redox and proteostasis program. Immediately after MOMP, oxTRX1 forms a disulfide with MIA40 Cys4 at the MIA40-AIFM1 interface. Subsequently, DPP8/9 cleave AK2, targeting this competing AIFM1 ligand for degradation. Biochemical reconstitution and cryo-EM reveal that oxTRX1 promotes AIFM1 dimerization and preferentially stabilizes MIA40-AIFM1 over AK2-AIFM1. Thus, redox-dependent complex assembly and selective proteolysis converge to favor MIA40-bound AIFM1 after MOMP. These findings suggest how post-MOMP remodeling could reinforce NLRP1 restraint during apoptosis and how disruption of this program could trigger pyroptosis.

cell biology↗

bioRxiv2023

The interaction between NLRP1 and oxidized TRX1 involves a transient disulfide bond

NLRP1 is an innate immune receptor that detects pathogen-associated signals, assembles into a multiprotein structure called an inflammasome, and triggers a proinflammatory form of cell death called pyroptosis. We previously discovered that the oxidized, but not the reduced, form of thioredoxin-1 directly binds to NLRP1 and represses inflammasome formation. However, the molecular basis for NLRP1s selective association with only the oxidized form of TRX1 has not yet been established. Here, we leveraged Alphafold-Multimer, site-directed mutagenesis, thiol-trapping experiments, and mass spectrometry to reveal that a specific cysteine residue (C427 in humans) on NLRP1 forms a transient disulfide bond with oxidized TRX1. Overall, this work demonstrates how NLRP1 monitors the cellular redox state, further illuminating an unexpected connection between the intracellular redox potential and the innate immune system.

immunology↗

bioRxiv2022

The NLRP1 and CARD8 inflammasomes detect reductive stress

The danger signals that activate the NLRP1 and CARD8 inflammasomes have not been fully established. We recently discovered that cytosolic peptide accumulation activates these inflammasomes. In addition, we found that the oxidized form of TRX1 binds to and represses NLRP1, suggesting that NLRP1 also detects a lack of reactive oxygen species, or reductive stress. However, no agents that induce reductive stress were known to test this premise. Here, we identify and characterize several radical-trapping antioxidants, including JSH-23, that induce reductive stress. We show that these compounds accelerate the proteasome-mediated degradation of the repressive N-terminal fragments of NLRP1 and CARD8, releasing the inflammasome-forming C-terminal fragments from autoinhibition. Moreover, we found that reductive stress and peptide accumulation together trigger much more intense inflammasome activation than either signal alone. Overall, this work validates chemical probes that induce reductive stress, and establishes reductive stress alongside peptide accumulation as the key inflammasome-activating danger signals.

immunology↗

bioRxiv2022

Cytosolic peptide accumulation activates the NLRP1 and CARD8 inflammasomes

NLRP1 and CARD8 are related sensors that form inflammasomes, but the danger signals that they detect are not fully established. These proteins undergo autoproteolysis, generating repressive N-terminal (NT) and inflammatory C-terminal (CT) fragments. The proteasome- mediated degradation of the NT releases the CT from autoinhibition, but the CT is then sequestered in a complex with the full-length sensor and DPP9. Here, we show that cytosolic peptide accumulation activates these inflammasomes. We found that a diverse array of peptides accelerates NT degradation, and those with N-terminal XP sequences also destabilize the ternary complexes. Peptides interfere with many biological processes, including protein folding. We show that unrelated agents that disrupt protein folding also induce NT degradation, but do not cause inflammasome activation because DPP9 sequesters the CT fragments in the absence of XP peptides. Overall, these results indicate that NLRP1 and CARD8 detect protein misfolding that is associated with peptide accumulation.

immunology↗

bioRxiv2022

A ubiquitin-independent proteasome pathway controls the CARD8 inflammasome

CARD8 is a pattern-recognition receptor that forms a caspase-1-activating inflammasome. CARD8 undergoes autoproteolysis, generating an N-terminal (NT) fragment with a disordered region and a ZU5 domain and a C-terminal (CT) fragment with UPA and CARD domains. DPP8 and DPP9 (DPP8/9) inhibitors, including Val-boroPro (VbP), accelerate the degradation of the NT fragment via a poorly characterized proteasome-mediated pathway, thereby releasing the inflammatory CT fragment from autoinhibition. Here, we show that the core 20S proteasome, which degrades disordered and misfolded proteins independent of ubiquitin, controls CARD8 activation. In unstressed cells, the 20S proteasome degrades just the NT disordered region, leaving behind the folded ZU5, UPA, and CARD domains to act as an inhibitor of inflammasome assembly. In VbP-stressed cells, the 20S proteasome degrades the entire NT fragment, perhaps due to ZU5 domain unfolding, freeing the CT fragment from autoinhibition. Overall, this work shows that CARD8 NTs susceptibility to 20S proteasome-mediated degradation controls inflammasome activation.

immunology↗

bioRxiv2021

Oxidized thioredoxin-1 restrains the NLRP1 inflammasome

At least six human proteins detect danger-associated signals, assemble into complexes called inflammasomes, and trigger pyroptotic cell death. NLRP1 was the first protein discovered to form an inflammasome, but the danger signals and molecular mechanisms that control its activation have not yet been fully established. Here, we report that the NACHT-LRR region of NLRP1 directly binds to oxidized form of thioredoxin-1 (TRX1). We found that NLRP1 requires the ATPase activity of its NACHT domain to associate with TRX1, and that this interaction represses inflammasome activation. Moreover, we discovered that several patient-derived missense mutations in the NACHT-LRR region of NLRP1 weaken TRX1 binding, leading to inflammasome hyperactivation and autoinflammatory disease. Overall, our results establish that oxidized TRX1 binds to and restrains the NLRP1 inflammasome, thereby revealing a link between the cellular redox environment and innate immunity.

immunology↗

bioRxiv2019

Human caspase-1 autoproteolysis is required for ASC-dependent and -independent inflammasome activation

Pathogen-related signals induce a number of cytosolic pattern-recognition receptors (PRRs) to form canonical inflammasomes, which activate pro-caspase-1 and trigger pyroptotic cell death. All well-studied PRRs oligomerize with the pro-caspase-1-adapter protein ASC to generate a single large structure in the cytosol, which induces the autoproteolysis and activation of the pro-caspase-1 zymogen. However, several PRRs can also directly interact with pro-caspase-1 without ASC, forming much smaller \"ASC-independent\" inflammasomes. It is currently thought that pro-caspase-1 autoproteolysis does not occur during, and is not required for, ASC-independent inflammasome activation. Here, we show that the related human PRRs NLRP1 and CARD8 exclusively form ASC-dependent and ASC-independent inflammasomes, respectively, identifying CARD8 as the first PRR that cannot form an ASC-containing signaling platform. Despite their different structures, we discovered that both the NLRP1 and CARD8 inflammasomes require pro-caspase-1 autoproteolysis between the small and large catalytic subunits to induce pyroptosis. Thus, pro-caspase-1 self-cleavage is an obligate regulatory step in the activation of human canonical inflammasomes.

immunology↗