Regulation of the NLRP3 inflammasome by autophagy and mitophagy

Abstract

The NLRP3 inflammasome is a multiprotein complex that upon activation by the innate immune system drives a broad inflammatory response. The primary initial mediators of this response are pro-IL-1β and pro-IL-18, both of which are in an inactive form. Formation and activation of the NLRP3 inflammasome activates caspase-1, which cleaves pro-IL-1β and pro-IL-18 and triggers the formation of gasdermin D pores. Gasdermin D pores allow for the secretion of active IL-1β and IL-18 initiating the organism-wide inflammatory response. The NLRP3 inflammasome response can be beneficial to the host; however, if the NLRP3 inflammasome is inappropriately activated it can lead to significant pathology. While the primary components of the NLRP3 inflammasome are known, the precise details of assembly and activation are less well defined and conflicting. Here, we discuss several of the proposed pathways of activation of the NLRP3 inflammasome. We examine the role of subcellular localization and the reciprocal regulation of the NLRP3 inflammasome by autophagy. We focus on the roles of mitochondria and mitophagy in activating and regulating the NLRP3 inflammasome. Finally, we detail the impact of pathologic NLRP3 responses in the development and manifestations of pulmonary disease.

Keywords: NLRP3; caspase‐1; inflammasome; lung injury; mitochondria.

Figure 1.. NLRP3 inflammasome activation.

Classic NLRP3 inflammasome activation is a two-step process. The priming signal, originating from pathogen-associated molecular patterns (PAMPs) such as LPS via toll-like receptors (TLRs) or cytokines like TNF, induces NF-κB-dependent transcription of NLRP3 and pro-IL-1β. The second signal, necessary for inflammasome formation, is provided by NLRP3 agonists (e.g., ATP, silica, MSU), that have been shown to act through numerous cellular signals, including K+ efflux, Ca2+ influx, mitochondrial dysfunction, reactive oxygen species (ROS), and mitochondrial DNA (mtDNA) release. Separately, RNA viruses activate NLRP3 via mitochondrial antiviral signaling protein (MAVS). Activated caspase-1 processes pro-IL-1β, pro-IL-18, and GSDMD into their active forms, with GSDMD forming membrane pores that can trigger pyroptotic cell death. In its inactive state, NLRP3 binds to mitochondria, interacting with MAVS, cardiolipin, and mitofusin-2. In a discrete model, PKD-mediated phosphorylation releases NLRP3 from the mitochondria-associated ER membrane (MAM), facilitating its assembly at the trans-Golgi network (TGN). NLRP3, upon activation by ion gradient disruptions, organelle dysfunction, and/or metabolic shifts, traffics along microtubules in a process dependent on HDAC6 and dynein. At the microtubule-organizing center (MTOC), NLRP3 associates with NEK7, leading to inflammasome assembly. Additionally, in human monocytes, an alternative activation pathway involves TLR4-induced RIPK1-, FADD-, and caspase-8-dependent activation of NLRP3, resulting in IL-1β secretion in response to LPS alone.

Figure 2.. Different types of autophagy in mammalian cells.

(A) In chaperone-mediated autophagy, the KFERQ motif of the target cytosolic protein is recognized by heat shock protein 70 (HSC70c) along with other chaperones which deliver the protein to the lysosome via lysosome-associated membrane protein type 2A (LAMP2A). (B) In microautophagy, there is either invagination of the lysosomal membrane followed by fission with Endosomal Sorting Complex Required for Transport (ESCRT), or extension of lysosomal membrane followed by fusion with phagophore-like structure for trapping cytosolic components. (C) In macroautophagy, cytosolic components including damaged organelles are sequestered in double membrane vesicles or autophagosomes which then fuse with lysosomes to form autolysosomes where degradation occurs.

Figure 3.. Mitophagy and NLRP3 inflammasome.

Mitochondrial damage can trigger the Parkin-PINK1-mediated mitophagy pathway, which involves the ubiquitination (Ub) of mitochondrial outer membrane (OMM) proteins and the recruitment of p62. Additionally, damaged mitochondria may expose the inner membrane phospholipid cardiolipin on their surface, recruiting LC3b and resulting in mitophagy. Both these pathways have the potential to activate the NLRP3 inflammasome, however this activation is inhibited in the presence of active caspase-1. Increased calcium efflux via mucolipin-1 (TRPML1) from lysosomes can exacerbate lysosomal and mitochondrial damage, promoting NLRP3 inflammasome formation. Furthermore, particulate NLRP3 agonists, such as silica, alum, and MSU crystals, can induce lysosomal damage or elevate reactive oxygen species (ROS), which also facilitates the formation of the NLRP3 inflammasome.