Iron catalysis of lipid peroxidation in ferroptosis: Regulated enzymatic or random free radical reaction?

Abstract

Duality of iron as an essential cofactor of many enzymatic metabolic processes and as a catalyst of poorly controlled redox-cycling reactions defines its possible biological beneficial and hazardous role in the body. In this review, we discuss these two "faces" of iron in a newly conceptualized program of regulated cell death, ferroptosis. Ferroptosis is a genetically programmed iron-dependent form of regulated cell death driven by enhanced lipid peroxidation and insufficient capacity of thiol-dependent mechanisms (glutathione peroxidase 4, GPX4) to eliminate hydroperoxy-lipids. We present arguments favoring the enzymatic mechanisms of ferroptotically engaged non-heme iron of 15-lipoxygenases (15-LOX) in complexes with phosphatidylethanolamine binding protein 1 (PEBP1) as a catalyst of highly selective and specific oxidation reactions of arachidonoyl- (AA) and adrenoyl-phosphatidylethanolamines (PE). We discuss possible role of iron chaperons as control mechanisms for guided iron delivery directly to their "protein clients" thus limiting non-enzymatic redox-cycling reactions. We also consider opportunities of loosely-bound iron to contribute to the production of pro-ferroptotic lipid oxidation products. Finally, we propose a two-stage iron-dependent mechanism for iron in ferroptosis by combining its catalytic role in the 15-LOX-driven production of 15-hydroperoxy-AA-PE (HOO-AA-PE) as well as possible involvement of loosely-bound iron in oxidative cleavage of HOO-AA-PE to oxidatively truncated electrophiles capable of attacking nucleophilic targets in yet to be identified proteins leading to cell demise.

Keywords: 15-lipoxygenase; Ferroptosis; GPX4; Glutathione; Hydroperoxy-arachidonoyl-phosphatidylethanolamine; Iron; Iron chaperons; Lipid peroxidation.

Figure 1

Three pillars of ferroptosis.

Figure 2

Schema of cellular iron homeostasis. The schema illustrates the various metalloproteins and metallochaperons involved in the influx of dietary iron into the cell, its import/transport into different organelles via different proteins, and its efflux from the cell. dCytB: Duodeneal cytochrome B, DMT1 (SLC11A2): Natural resistance-associated macrophage protein 2, Fe-L_b: Low molecular mass iron complexes with cellular ligands, FE-S: Iron-Sulphur clusters, Ferritin: Protein that stores Iron, Fpn (SLC40A1): Ferroportein-1 iron transporter, Ferric reductase: catalyses reduction of Fe(III) to Fe(II), Ferroxidase: catalyses oxidation of Fe^(II) to Fe^(III), GSH: Reduced Glutathione, HEME: Heme, NCOA4: Nuclear receptor coactivator 4, Autophagic cargo receptor for ferritin., LOX: Lipoxygenase, AA-OOH; Hydroperoxy-arachidonic acid, HOOH; hydrogen peroxide; PCBP1/2: Poly(rC)-binding protein-1, Iron chaperone, PE: Phosphatidylethanolamine lipid,Steap: Metalloreductase, TF: Transferrin, Tfr1: Transferrin Receptor protein 1, ZIP14/8 (SCL39A14): Zinc and iron permease.

Figure 3

Schema illustrating three pillars of ferroptotic program:Lipid peroxidation, Iron handling pathways and GPX4-dependent reduction of hydroperoxy-phospholipids. ACSL, Acyl-CoA synthase; LCAT, Lyso-phospholipid acyl transferase; PEBP1, Phosphatidylethanolamine binding protein 1; LOX, Lipoxygenase; GSH, Glutathione; GS-SG, Oxidized glutathione; AA, arachidonic acid; PE-AA-OOH, Hydroperoxy-arachidonoyl phosphatidylethanolamine; PE-AA-OH, Hydroxy-arachidonoyl phosphatidylethanolamine;GPX4, Glutathione peroxidase 4; AH, Antioxidants (phenols, aromatic amines); Fe-L_b, Low molecular mass iron complexes with cellular ligands; TF, Transferrin; Tfr, Transferrin receptor.

Figure 4

Assessment of oxygenated phospholipid species in mouse liver. Typical MS spectra of phosphatidylethanolamine (PE) (A-a) and phosphatidylcholine (PC) (B-a) obtained from WT mouse liver. Analysis of oxygenated PE species (A-b) and oxygenated PC (B-b) in liver of PCBP1-deleted (KO) vs. wild type (WT) mice. Male mice were 5–6 weeks old and maintained on a synthetic, defined diet containing adequate, but not elevated, amounts of iron (50 ppm). Ferroptotic cell death signals (hydroperoxy-PE species) are shown as closed diamonds. N=6.

Scheme 1

Inhibition of GPX4 leads to the formation of alkoxyl radicals.

Scheme 2

Proposed reaction sequence leading to ferroptosis. Reactions with two arrows denote multiple steps that are omitted from the reaction scheme.