Oxidized thioredoxin-1 restrains the NLRP1 inflammasome

Abstract

The danger signals that activate the NLRP1 inflammasome have not been established. Here, we report that the oxidized, but not the reduced, form of thioredoxin-1 (TRX1) binds to NLRP1. We found that oxidized TRX1 associates with the NACHT-LRR region of NLRP1 in an ATP-dependent process, forming a stable complex that restrains inflammasome activation. Consistent with these findings, patient-derived and ATPase-inactivating mutations in the NACHT-LRR region that cause hyperactive inflammasome formation interfere with TRX1 binding. Overall, this work strongly suggests that reductive stress, the cellular perturbation that will eliminate oxidized TRX1 and abrogate the TRX1-NLRP1 interaction, is a danger signal that activates the NLRP1 inflammasome.

The NLRP1 and CARD8 inflammasomes detect reductive stress

“… 21 , 28 MeBs, which synergizes with VbP to induce more pyroptosis in these cell lines, was used as a positive control in this experiment. 8 , 26 Consistent with our previous results, 27 we found that dithiothreitol (DTT), N-acetyl cysteine (NAC), reduced L-glutathione (GSH), and trolox did not synergize with VbP to induce more NLRP1 or CARD8 activation in this assay ( Figures 1A , 1B , and S2A ). Intriguingly, we observed that ferrostatin-1 (Fer-1), a lipophilic antioxidant that blocks an iron-dependent form of cell death called ferroptosis, 29 unlike other commonly used antioxidants, appeared to slightly synergize with VbP in RAW 264.7 and MV4;11 cells at a high (40 μM) concentration ( Figures 1A , 1B , and S2A ).…”

“…Indeed, oxidative power is critical for protein folding in the ER 44 ; it seems possible that oxidative power plays a similar, but as-yet-unknown, role in the folding of cytosolic proteins, including NLRP1’s and CARD8’s NT fragments. In addition, we recently discovered that oxidized TRX1 stabilizes NLRP1 NT’s structure, 27 which we now speculate serves as a “checkpoint” to ensure misfolding is indeed associated with reductive stress. Whether a redox-sensitive protein similarly regulates the folding of the CARD8 NT remains to be determined.…”

“…Briefly, NLRP1 has NT nucleotide-binding (NACHT) and leucine-rich (LRR) domains ( Figure S1 ), and these domains associate with the oxidized form of thioredoxin-1 (TRX1). 27 We found that genetic loss of TRX1 sensitizes cells to VbP-induced pyroptosis, showing that oxidized TRX1 restrains NLRP1 activation. These results predicted that agents that deplete ROS and reduce TRX1 would synergize with VbP to induce more NLRP1-dependent pyroptosis.…”

“…We recently discovered that the oxidized, but not the reduced, form of TRX1 binds to and represses NLRP1. 27 As such, we predicted that reductive stress would abrogate this interaction and lead to inflammasome activation. However, no agents were known that induce reductive stress to test this hypothesis.…”

“…In addition, it is not yet known whether reductive stress, protein folding, and XP peptide accumulation are all in some way directly related ( Figure S1B ). 27 …”