Ferroptosis turns 10: Emerging mechanisms, physiological functions, and therapeutic applications

Abstract

Ferroptosis, a form of cell death driven by iron-dependent lipid peroxidation, was identified as a distinct phenomenon and named a decade ago. Ferroptosis has been implicated in a broad set of biological contexts, from development to aging, immunity, and cancer. This review describes key regulators of this form of cell death within a framework of metabolism, ROS biology, and iron biology. Key concepts and major unanswered questions in the ferroptosis field are highlighted. The next decade promises to yield further breakthroughs in the mechanisms governing ferroptosis and additional ways of harnessing ferroptosis for therapeutic benefit.

Figure 1.. Key milestones and growth in the literature of ferroptosis over time.

The number of publications listed in PubMed in each year is indicated, as well as key discoveries related to ferroptosis in each year. A glossary of terms related to ferroptosis is also shown.

Figure 2.. Mechanisms of ferroptosis.

Protein names are in black, those that facilitate ferroptosis are in red circles, and those that suppress ferroptosis in blue circles. Mechanisms are in rectangles. Small molecule inhibitors, lipids and metabolite structures are shown and labeled in green for those that suppress ferroptosis and red for those that induce ferroptosis, and purple for those that neither promote not suppress ferroptosis. The bottom right membrane is the plasma membrane, where system x_c⁻ imports cystine, which is reduced in the cell to the amino acid cysteine. Cysteine and glutamate are used in the biosynthesis of reduced glutathione, which is in turn used by GPX4 to reduce reactive PUFA phospholipid hydroperoxides (PUFA-PL-OOH) to non-reactive and non-lethal PUFA phospholipid alcohols (PUFA-PL-OH). Alternatively, the oxidized PUFA-OOH tail can be cleaved from a phospholipid by the action of iPLA2β, suppressing death. The membrane in the middle represents cellular membranes that experience lipid peroxidation, such as the ER. PUFA-PLs are oxidized by labile Fe(II) and Fe(II)-dependent enzymes, such as ALOXs in conjunction with PEBP1, and POR. Fe(III) is imported into cells by Tf through TfR1, after which it is reduced to Fe(II) and imported via DMT1. Iron is stored as Fe(III) in ferritin, where it is not available to promote ferroptosis. Export of iron in MVBs is promoted by prom2, which suppresses ferroptosis. In the top panel, acetyl CoA is used to make free PUFAs, which are activated by ACSL4, LPCAT3 and ACSL1 to generate PUFA-PLs; ACSL4 can be phosphorylated by PKCβII to further activate it. The yellow box shows the process of generating PUFA-PLs and the green box shows how PUFA-PLs are oxidized. Abbreviations: ALOXs, lipoxygenases; AMPK, adenosine-monophosphate-activated protein kinase; ACC, acetyl coenzyme A carboxylase; ACSL, acyl-CoA synthetase long chain family member; ATM, ATM serine/threonine kinase; BH₄, tetrahydrobiopterin; CDO1, Cysteine dioxygenase type 1; CoA, coenzyme A; CoQ₁₀, coenzyme Q₁₀; cys, cysteine; DHODH, dihydroorotate dehydrogenase (quinone); DMT1, ferrous ion membrane transport protein DMT1; FSP, ferroptosis suppressor protein 1/ AIFM2; GCH1, GTP cyclohydrolase 1; GCLC, glutamate-cysteine ligase catalytic subunit; Glu, glutamate; GPX4, glutathione peroxidase 4; GSH, glutathione; IL4i1, interleukin-4-induced 1; In3Py, indole-3-pyruvate; iPLA2β, phospholipase A2 group VI; LPCAT3, lysophosphatidylcholine acyltransferase 3; lysoPL, lysophospholipid; MDR1, ATP binding cassette subfamily B member 1/ABCB1; MUFA, monounsaturated fatty acid; MVB, multivesicular body; NADPH, reduced nicotinamide adenine dinucleotide phosphate; PEBP1, phosphatidylethanolamine binding protein 1; PKCβII, protein kinase C beta type isoform 2; PL, phospholipid; POR, Cytochrome p450 oxidoreductase; prom2, prominin-2; PUFA, polyunsaturated fatty acid; PUFA-PL-OOH, phospholipid with peroxidized polyunsaturated fatty acyl tail; ROS, reactive oxygen species; system x_c⁻, sodium-independent, anionic amino acid transport system; Tf, transferrin; TfR1, transferrin receptor protein 1; YAP, Yes1 associated transcriptional regulator

Figure 3.. Role of organelles and organs in ferroptosis.

(A) The contributions of major organelles in ferroptosis are shown. (B) Ferroptosis has been implicated in a variety of diseases, organs and tissues, as shown. Abbreviations: AD, Alzheimer’s Disease; ALS, amyotrophic lateral sclerosis; HD, Huntington Disease; PD, Parkinson’s Disease; PM, particulate matter; SARS-CoV2, severe acute respiratory syndrome, coronavirus 2; SSMD, Sedaghatian-type Spondylometaphyseal Dysplasia

Figure 4.. Physiological functions of ferroptosis.

(A) Anti-viral immunity is promoted by selenium supplementation. A selenium-rich diet enhances expression of the selenoprotein GPX4, which suppresses ferroptosis in CD4⁺ T_FH cells, which in turn promotes increased memory B cells and long-lasting viral immunity. (B) Role of ferroptosis in tumor suppression. Tumor suppressors are indicated in orange boxes, proteins that suppress ferroptosis are in blue circles and those that promote ferroptosis are in red circles. Small molecules that suppress ferroptosis are in light blue circles. A PUFA-rich diet promotes production of PUFA-PLs, which promote tumor ferroptosis. CD8⁺ T cells further promote tumor cell ferroptosis by releasing IFNγ and AA. A cholesterol-rich diet promotes increased expression of CF36 on CD8+ T cells, which caused them to take up PUFAs themselves, leading to their death by ferroptosis and promoting tumor formation. (C) Ferroptosis increases with aging. Rats and mice experience increased ferroptosis markers as they age. Nematodes (C. elegans) experience increased iron content and depletion of glutathione as they age, leading to increased ferroptosis. (D) Ferroptosis is involved in development. Nucleated erythrocytes undergo ferroptosis prior to enucleation and erythrocyte maturation. A fungal pathogen has evidence of ferroptosis during its maturation required for plant infection. Abbreviations: AA, arachidonic acid; ALOXs, lipoxygenases; BAP1, ubiquitin carboxyl-terminal hydrolase; CoA, coenzyme A; GPX4, glutathione peroxidase 4; IFNγ, interferon gamma; MLL4, histone-lysine N-methyltransferase 2B; NFS1, cysteine desulfurase, mitochondrial; p53, p53 tumor suppressor protein; PUFA, polyunsaturated fatty acid; PUFA-PL, phospholipid with polyunsaturated fatty acid tail; SCD1, stearoyl-coenzyme A desaturase 1; SLC7A11, solute carrier family 7 member 11; T_FH, Follicular helper T cells

Figure 5.. Markers of ferroptosis.

Four classes of markers for detecting ferroptotic cells are shown, along with recommendations for their use. The table on the bottom compares how these markers may appear in various contexts: during ferroptosis, apoptosis, necroptosis, oxidative stress, iron stress, or mitochondrial stress. While some markers may be activated by these other types of stress, if at least three markers are used, or a suitable combination of two markers (such as lipid peroxidation and TfR1 mobilization) ferroptosis can be distinguished from these other stress conditions.

Figure 6.. Key concepts in ferroptosis.

Six key concepts that aid in understanding ferroptosis are described. A. Different lipids have different effects on ferroptosis. Lipids with saturated fatty acids generally don’t affect ferroptosis, whereas those with monounsaturated fatty acids suppress ferroptosis and those with polyunsaturated fatty acids promote ferroptosis. (B) Reactive oxygen species (ROS) can occur without inducing ferroptosis. In some cases, soluble ROS accumulate and cause oxidative stress without inducing lipid peroxidation or ferroptosis. (C) Iron can induce effects unrelated to ferroptosis, and the oxidation state of iron is critical for ferroptosis. Some iron toxicity does not depend on lipid peroxidation. The ability of iron to promote ferroptosis mostly depends on it being in the Fe(II) state, although lipoxygenase generally require iron to be in Fe(III) to be activated. (D) The degree to which ferroptosis is inflammatory is unclear. Ferroptosis involves the release of some factors, which can be inflammatory, but perhaps not to the same extent as during classic necrosis or pyroptosis. (E) The relationship of ferroptosis to necrosis is unclear. Whether necrosis is a broader umbrella or a distinct process remains uncertain. (F) Metabolism, iron regulation, and ROS defenses together control ferroptosis. Free polyunsaturated fatty acids can be obtained through biosynthesis or diet, and are converted into membrane-localized phospholipids. Iron promotes oxidation of PUFA-PLs to PUFA-PL hydroperoxides, which can be eliminated through reduction by glutathione peroxidase 4, using cysteine-derived glutathione, yielding non-toxic lipid alcohols, or by iPLA2β, yielding free oxidized PUFA and lysolipids, which undergo further lipid remodeling. Oxidized PUFA-PLs can propagate in the presence of Fe(II), eventually yielding membrane damage and/or generation of reactive lipid-derived electrophiles. CoQ₁₀ and BH₄ and the pathways that generate them can block propagation of oxidized PUFA-PLs. Metabolism-related pathways are shown in red, iron regulation pathways are shown in green, and ROS defense pathways are shown in blue. Abbreviations: BH₄, tetrahydrobiopterin; CoA, coenzyme A; CoQ₁₀, coenzyme Q₁₀; cys, cysteine; DMT1, ferrous ion membrane transport protein DMT1; GPX4, glutathione peroxidase 4; iPLA2β, phospholipase A2 group VI; lysoPL, lysophospholipid; MUFA, monounsaturated fatty acid; MVB, multivesicular body; NADPH, reduced nicotinamide adenine dinucleotide phosphate; PL, phospholipid; prom2, prominin-2; PUFA, polyunsaturated fatty acid; PUFA-PL, phospholipid containing a polyunsaturated fatty acyl lipid tail; PUFA-PL-OOH, phospholipid with peroxidized polyunsaturated fatty acyl tail; ROS, reactive oxygen species; system x_c⁻, sodium-independent, anionic amino acid transport system; Tf, transferrin; TfR1, transferrin receptor protein 1.