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.

Background and Aims Iron is essential yet also highly chemically reactive and potentially toxic. The mechanisms that allow cells to use iron safely are not clear; defects in iron management are a causative factor in the cell‐death pathway known as ferroptosis. Poly rC binding protein 1 (PCBP1) is a multifunctional protein that serves as a cytosolic iron chaperone, binding and transferring iron to recipient proteins in mammalian cells. Although PCBP1 distributes iron in cells, its role in managing iron in mammalian tissues remains open for study. The liver is highly specialized for iron uptake, utilization, storage, and secretion. Approach and Results Mice lacking PCBP1 in hepatocytes exhibited defects in liver iron homeostasis with low levels of liver iron, reduced activity of iron enzymes, and misregulation of the cell‐autonomous iron regulatory system. These mice spontaneously developed liver disease with hepatic steatosis, inflammation, and degeneration. Transcriptome analysis indicated activation of lipid biosynthetic and oxidative‐stress response pathways, including the antiferroptotic mediator, glutathione peroxidase type 4. Although PCBP1‐deleted livers were iron deficient, dietary iron supplementation did not prevent steatosis; instead, dietary iron restriction and antioxidant therapy with vitamin E prevented liver disease. PCBP1‐deleted hepatocytes exhibited increased labile iron and production of reactive oxygen species (ROS), were hypersensitive to iron and pro‐oxidants, and accumulated oxidatively damaged lipids because of the reactivity of unchaperoned iron. Conclusions Unchaperoned iron in PCBP1‐deleted mouse hepatocytes leads to production of ROS, resulting in lipid peroxidation (LPO) and steatosis in the absence of iron overload. The iron chaperone activity of PCBP1 is therefore critical for limiting the toxicity of cytosolic iron and may be a key factor in preventing the LPO that triggers the ferroptotic cell‐death pathway.

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Bipolar disorder (BD), which is characterized by depression and mania, affects 1-2% of the world population. Current treatments are effective in only 40-60% of cases and cause severe side effects. Valproate (VPA) is one of the most widely used drugs for the treatment of BD, but the therapeutic mechanism of action of this drug is not understood. This knowledge gap has hampered the development of effective treatments. To identify candidate pathways affected by VPA, we performed a genome-wide expression analysis in yeast cells grown in the presence or absence of the drug. VPA caused up-regulation of FEN1 and SUR4, encoding fatty acid elongases that catalyze the synthesis of very long chain fatty acids (C24 to C26) required for ceramide synthesis. Interestingly, fen1Δ and sur4Δ mutants exhibited VPA sensitivity. In agreement with increased fatty acid elongase gene expression, VPA increased levels of phytoceramide, especially those containing C24-C26 fatty acids. Consistent with an increase in ceramide, VPA decreased the expression of amino acid transporters, increased the expression of ER chaperones, and activated the unfolded protein response element (UPRE), suggesting that VPA induces the UPR pathway. These effects were rescued by supplementation of inositol and similarly observed in inositol-starved ino1Δ cells. Starvation of ino1Δ cells increased expression of FEN1 and SUR4, increased ceramide levels, decreased expression of nutrient transporters, and induced the UPR. These findings suggest that VPA-mediated inositol depletion induces the UPR by increasing the de novo synthesis of ceramide.

Poly(rC)-binding protein (PCBP1) is a multifunctional adaptor protein that can coordinate single-stranded nucleic acids and iron–glutathione complexes, altering the processing and transfer of these ligands through interactions with other proteins. Multiple phenotypes are ascribed to cells lacking PCBP1, but the relative contribution of RNA, DNA, or iron chaperone activity is not consistently clear. Here, we report the identification of amino acid residues required for iron coordination on each structural domain of PCBP1 and confirm the requirement of iron coordination for binding target proteins BolA2 and ferritin. We further construct PCBP1 variants that lack either nucleic acid– or iron-binding activity and examine their functions in human cells and mouse tissues depleted of endogenous PCBP1. We find that these activities are separable and independently confer essential functions. While iron chaperone activity controls cell cycle progression and suppression of DNA damage, RNA/DNA-binding activity maintains cell viability in both cultured cell and mouse models. The coevolution of RNA/DNA binding and iron chaperone activities on a single protein may prove advantageous for nucleic acid processing that depends on enzymes with iron cofactors.

the widely used mood stabilizer valproate (VpA) causes perturbation of energy metabolism, which is implicated in both the therapeutic mechanism of action of the drug as well as drug toxicity. to gain insight into these mechanisms, we determined the effects of VPA on energy metabolism in yeast. VpA treatment increased levels of glycolytic intermediates, increased expression of glycolysis genes, and increased ethanol production. increased glycolysis was likely a response to perturbation of mitochondrial function, as reflected in decreased membrane potential and oxygen consumption. interestingly, yeast, mouse liver, and isolated bovine cytochrome c oxidase were directly inhibited by the drug, while activities of other oxidative phosphorylation complexes (III and V) were not affected. These findings have implications for mechanisms of therapeutic action and toxicity. Bipolar disorder (BD) is a severe psychiatric illness characterized by shifts in mood, ranging from mania to depression. It affects at least 1% of the population and leads to suicide in 15% of cases 1. BD patients exhibit a higher prevalence of obesity, cardiovascular disease, and diabetes than the general population 2,3. Many studies have shown that the pathophysiology of BD involves altered energy metabolism 4-8. While most of the metabolic markers measured indicate mitochondrial dysfunction in BD 9-12 , some studies have suggested the presence of increased mitochondrial activity in the manic phase of BD 13 , including increased energy generation 14 , basal metabolic rate 15 , uric acid 16 , and calcium ions 17. Valproate (VPA) is one of several mood stabilizers approved by the FDA for the treatment of BD 18,19 , epilepsy 20,21 , and migraine 22. The mechanism of action of VPA is not understood 23. VPA is effective in only 40-60% of cases and can cause serious side effects, including hepatotoxicity and teratogenicity 24. Hepatotoxicity can be life-threatening 25,26 and may occur even at therapeutic doses 27. Although rare, lethal hepatotoxicity associated with VPA has been described in both children 28 and adults 29. The prominent feature of this type of hepatotoxicity is microvesicular steatosis 30 , consistent with mitochondrial dysfunction 31. In agreement with this, patients with congenital defects in mitochondrial metabolism are at a higher risk for susceptibility to VPA toxicity 32-34. VPA exerts numerous documented effects on mitochondrial metabolism. It is metabolized by and inhibits ß-oxidation through several mechanisms 35. VPA and its metabolites sequester coenzyme A (CoA), depleting mitochondrial CoA 36. Furthermore, studies suggest that both unesterified VPA as well as VPA acyl-CoA esters inhibit fatty acid oxidation enzymes 37,38. In addition to affecting ß-oxidation, VPA inhibits α-ketoglutarate dehydrogenase, a key enzyme of the tricarboxylic acid (TCA) cycle 39,40. Inhibition of this enzyme is a proposed mechanism underlying decreased TCA cycle flux in the presence of VPA 41. VPA also decreases levels of carnitine 42,43 , which transpo...

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Bipolar disorder (BD) is a severe psychiatric illness affecting ∼1% of the world population. Valproate (VPA) and lithium, widely used for the treatment of BD, are not universally effective. These drugs have been shown to cause inositol depletion, but translating this observation to a specific therapeutic mechanism has been difficult, hampering the development of more effective therapies. We have shown previously in yeast that chronic VPA treatment induces the unfolded protein response due to increasing ceramide levels. To gain insight into the mechanisms activated during acute VPA treatment, we performed a genome-wide expression study in yeast treated with VPA for 30 min. We observed increased mRNA and protein levels of , which encodes an exporter of long chain bases dihydrosphingosine (DHS) and phytosphingosine (PHS), and further saw that VPA increased sensitivity of anΔ mutant to PHS, suggesting that VPA increases long chain base levels. Consistent with this, PHS levels were elevated in wild type and, to a greater extent, in Δ cells. Expression of genes (negative regulators of PHS synthesis) and of fatty acid elongase genes and were decreased, and expression of (exporter of PHS-1P) and (lyase that degrades DHS-1P and PHS-1P) was increased. These effects were more pronounced in medium lacking inositol, and were mirrored by inositol starvation of an Δ mutant. These findings provide a metabolic explanation as to how VPA-mediated inositol depletion causes increased synthesis of PHS and further support the therapeutic relevance of inositol depletion as a bipolar disorder treatment.