Structural basis for distinct inflammasome complex assembly by human NLRP1 and CARD8

Abstract

Nod-like receptor (NLR) proteins activate pyroptotic cell death and IL-1 driven inflammation by assembling and activating the inflammasome complex. Closely related sensor proteins NLRP1 and CARD8 undergo unique auto-proteolysis-dependent activation and are implicated in auto-inflammatory diseases; however, their mechanisms of activation are not understood. Here we report the structural basis of how the activating domains (FIINDUPA-CARD) of NLRP1 and CARD8 self-oligomerize to assemble distinct inflammasome complexes. Recombinant FIINDUPA-CARD of NLRP1 forms a two-layered filament, with an inner core of oligomerized CARD surrounded by an outer ring of FIINDUPA. Biochemically, self-assembled NLRP1-CARD filaments are sufficient to drive ASC speck formation in cultured human cells—a process that is greatly enhanced by NLRP1-FIINDUPA which forms oligomers in vitro. The cryo-EM structures of NLRP1-CARD and CARD8-CARD filaments, solved here at 3.7 Å, uncover unique structural features that enable NLRP1 and CARD8 to discriminate between ASC and pro-caspase-1. In summary, our findings provide structural insight into the mechanisms of activation for human NLRP1 and CARD8 and reveal how highly specific signaling can be achieved by heterotypic CARD interactions within the inflammasome complexes.

The role of inflammasomes in vascular cognitive impairment

“…The NLR-pyrin domain containing 1 (NLRP1) belongs to the NLR family of PRRs, with only a single variant of NLRP1 found in humans [ 194 , 219 ]. The NLRP1 receptor consists of an N-terminal PYD, NACHT, LRR, function-to-find domain (FIIND) and a C-terminal CARD domain.…”

“…The NLRP1 receptor consists of an N-terminal PYD, NACHT, LRR, function-to-find domain (FIIND) and a C-terminal CARD domain. The FIIND domain is unique to NLRP1 in the NLR family, and may play an autoinhibitory role [ 219 , 220 ] (Fig. 3 ).…”

“…NLRP1 oligomers appear to interact with ASC via homotypic PYD-PYD interactions [ 220 ]. After ASC speck formation, the ASC speck can recruit multiple effector inflammatory caspases via homotypic CARD-CARD interactions [ 219 ]. Despite being able to mediate ASC-independent NLRP1 inflammasome activation, NLRP1 activity level is higher in the presence of an ASC speck [ 220 ] (Fig.…”

Cryo-electron tomography of NLRP3-activated ASC complexes reveals organelle co-localization

“…The PYD and CARD of ASC are both required for punctum formation and signalling 15,17,18 . Structural studies on purified ASC PYD and ASC CARD show that both domains have a death fold and independently form helical filaments [19][20][21][22][23][24] . In the cell, fluorescence microscopy imaging suggests that PYD-PYD interactions drive ASC filament formation whereas CARD-CARD interactions promote cross-linking and compaction of PYD filaments into puncta 17,18,[23][24][25][26] .…”

“…The copyright holder for this preprint this version posted September 20, 2021. ; https://doi.org/10.1101/2021.09.20.461078 doi: bioRxiv preprint domains have a death fold and independently form helical filaments [19][20][21][22][23][24] . In the cell, fluorescence microscopy imaging suggests that PYD-PYD interactions drive ASC filament formation whereas CARD-CARD interactions promote cross-linking and compaction of PYD filaments into puncta 17,18,[23][24][25][26] .…”