Partners in Crime: The Interplay of Proteins and Membranes in Regulated Necrosis

Abstract

Pyroptosis, necroptosis, and ferroptosis are well-characterized forms of regulated necrosis that have been associated with human diseases. During regulated necrosis, plasma membrane damage facilitates the movement of ions and molecules across the bilayer, which finally leads to cell lysis and release of intracellular content. Therefore, these types of cell death have an inflammatory phenotype. Each type of regulated necrosis is mediated by a defined machinery comprising protein and lipid molecules. Here, we discuss how the interaction and reshaping of these cellular components are essential and distinctive processes during pyroptosis, necroptosis, and ferroptosis. We point out that although the plasma membrane is the common target in regulated necrosis, different mechanisms of permeabilization have emerged depending on the cell death form. Pore formation by gasdermins (GSDMs) is a hallmark of pyroptosis, while mixed lineage kinase domain-like (MLKL) protein facilitates membrane permeabilization in necroptosis, and phospholipid peroxidation leads to membrane damage in ferroptosis. This diverse repertoire of mechanisms leading to membrane permeabilization contributes to define the specific inflammatory and immunological outcome of each type of regulated necrosis. Current efforts are focused on new therapies that target critical protein and lipid molecules on these pathways to fight human pathologies associated with inflammation.

Keywords: inflammation; membrane permeabilization; pores; protein-lipid interactions; regulated necrosis.

Figure 1

Pyroptosis is a caspase-dependent form of regulated necrosis that critically depends on the pore forming activity of the gasdermins (GSDMs). Two different pathways can trigger pyroptosis. In the canonical one, external or endogenous danger signals are sensed by the inflammasome through pattern recognition receptors that interact with procaspase-1 either directly through a PYRIN-PAAD-DAPIN (PYD)/C-terminal caspase-recruitment domain (CARD) (PYD/CARD) domain or indirectly via an Apoptosis-associated speck-like protein containing a CARD (ASC) adaptor. The yellow ball represents the protein domain removed during the activation of caspase 1. Active caspase 1 cleaves and activates the inflammatory cytokines IL-1β and IL-18. In the non-canonical pathway, lipopolysaccharide (LPS) from Gram-negative bacteria binds to the CARD domain of caspase 4, 5, and 11. In both pathways, caspases mediate the cleavage of GSDMD, which allows the release of the N-terminal domain (GSDMD-N) from auto-inhibition by the C-domain (represented by a green ball). GSDMD-N translocates to the inner leaflet of the plasma membrane, where it interacts with anionic phospholipidsand oligomerizes forming arc or slits structures that evolve to a ring of a β-barrel pore. Blue balls are the polar head groups of membrane phospholipids.

Figure 2

Necroptosis is a caspase-independent form of regulated necrosis that critically depends on membrane permeabilization of mixed lineage kinase domain-like (MLKL) protein. Left: Necroptosis is initiated by various stimuli that all result in the formation of the necrosome, phosphorylation of MLKL, and its subsequent translocation to the plasma membrane. MLKL oligomerization is also modulated by its interaction with highly phosphorylated inositol phosphates (IPs), which are soluble products of lipid metabolism. MLKL activation facilitates its weak interaction with phosphatidylinositol phosphates (PIPs) at the membrane. Membrane binding promotes the exposure of new high-affinity sites and strong membrane interaction. Right: Different models of MLKL-mediated membrane permeabilization in necroptosis: (1) indirect activation of endogenous ion channels, (2) partial insertion in the bilayer forming amyloid fibers, and (3) formation of selective channels or pores.

Figure 3

Ferroptosis is a caspase-independent form of regulated necrosis that involves the production of phospholipid peroxides in cellular membranes. Left: Loss of glutathione peroxidase 4 (GPX4) activity is the main trigger of ferroptosis. Ras-selective Lethal Small Molecule 3 (RSL3), which directly inhibits GPX4, and erastin-1, which affects the catalytic cycle of GPX4 by perturbing the levels of glutathione (GSH), both trigger ferroptosis. Ferroptosis suppressor protein 1 (FSP1) and GTP cyclohydrolase-1 (GCH1) that target ubiquinone in the plasma membrane can regulate accumulation of membrane oxidized PUFAs-phospholipids. Acyl-CoA synthetase long-chain family 4 (ACSL4) and lysophosphatidylcholine acyltransferase 3 (LPCAT3) mediate the incorporation of PUFAs into membrane. Lipoxigenases (LOX) catalyzes the incorporation of oxygen into the PUFAs, to produce lipid hydroperoxides, which are accumulated at the membrane in the absence of GPX4 activity. Right: Different models of plasma membrane permeabilization in ferroptosis: induction of membrane curvature by the accumulation of phospholipid hydroperoxides (1) (2) alteration of the function of membrane proteins and/or stabilization of membrane curvature by endogenous proteins.