Assembly and regulation of ASC specks

Abstract

The inflammasome adapter ASC links activated inflammasome sensors to the effector molecule pro-caspase-1. Recruitment of pro-caspase-1 to ASC promotes the autocatalytic activation of caspase-1, which leads to the release of pro-inflammatory cytokines, such as IL-1β. Upon triggering of inflammasome sensors, ASC assembles into large helical fibrils that interact with each other serving as a supramolecular signaling platform termed the ASC speck. Alternative splicing, post-translational modifications of ASC, as well as interaction with other proteins can perturb ASC function. In several inflammatory diseases, ASC specks can be found in the extracellular space and its presence correlates with poor prognosis. Here, we review the role of ASC in inflammation, and focus on the structural mechanisms that lead to ASC speck formation, the regulation of ASC function during inflammasome assembly, and the importance of ASC specks in disease.

Keywords: CARD; COP; Fibril; POP; PTM; PYD; Prionoid; Speck.

Fig. 1

NMR structure of ASC (PDB ID: 2KN6) [33]. a Cartoon diagram of ASC, showing the PYD, the flexible linker and the CARD. b Cartoon diagram of the typical DD fold of the ASC–PYD. Indicated are the six α-helices (α1 to α6)

Fig. 2

ASC speck structure at different levels of complexity. a Confocal laser scanning microscopy image of a pyroptotic macrophage that expresses ASC-mCherry and has assembled an ASC speck (red). The cell was counterstained for nucleus (blue) and membranes (green). b High-resolution stimulated emission depletion (STED) image of an ASC speck. It shows distinct ASC fibrils emerging from the condensed, tightly cross-linked core of the ASC speck. c, d Schematic view of an ASC speck, containing the inflammasome sensor NLRP3 (green), which clusters and forms a seed for the prion-like polymerization of ASC into fibrils through homotypic interactions of its PYD (blue). The CARDs of ASC (red) form clusters on the surface of the PYD fibrils. These CARD clusters can either interact with other ASC–CARDs and thereby crosslink the ASC fibrils into a condensed speck, or they can form a seed for the assembly of polymeric CARD fibrils of pro-caspase-1. After polymerization of pro-caspase-1, the close proximity of several caspases leads to self-cleavage into the mature caspase-1, which is released from the speck. e Schematic view of the three-start ASC–PYD helix, indicating the direction of further polymerization and the three strands. f Detailed scheme of the three interfaces that mediate ASC–PYD interactions within the ASC fibril. The intra-strand interface I contributes the most to ASC PYD–PYD interactions, followed by the smaller inter-strand interfaces II and III

Fig. 3

ASC mRNA is alternatively spliced into four different isoforms. PYCARD consists of three exons. Exon 1 encodes the PYD, exon 2 encodes the flexible linker, and exon 3 encodes the CARD. ASC-a represents the canonical sequence, and contains all three exons, ASC-b lacks exon 2, ASC-c lacks 60 aa within the PYD and is the product of cryptic splice sides within exon 1, ASC-d is derived by a frame shift after the first 35 aa of the PYD, which results in a completely new exon 2 (exon 2′)

Fig. 4

Regulation of ASC speck assembly and function. a Pyrin only proteins (POPs) act as decoy-binding partners for PYD domains. They can either block ASC–PYD–PYD interactions or interactions between ASC and specific inflammasome sensors. The splice variants ASC-b and ASC-c as well as the CARD-only proteins (COPs) block different CARD–CARD interactions. b Post-translational modifications that regulate ASC function. Phosphorylation (P) and ubiquitination (Ub) that activate or inhibit ASC function are depicted. The respective kinase or the E3 ubiquitin ligase is indicated for every modification, as well as the inflammasome sensors involved in the process