2016 review

Regulators of Iron Homeostasis: New Players in Metabolism, Cell Death, and Disease

[Review of ]. TRENDS IN BIOCHEMICAL SCIENCES, 41(3), 274–286.

Source: Web Of Science
Added: August 6, 2018

Iron is necessary for life, but can also cause cell death. Accordingly, cells evolved a robust, tightly regulated suite of genes for maintaining iron homeostasis. Previous mechanistic studies on iron homeostasis have granted insight into the role of iron in human health and disease. We highlight new regulators of iron metabolism, including iron-trafficking proteins [solute carrier family 39, SLC39, also known as ZRT/IRT-like protein, ZIP; and poly-(rC)-binding protein, PCBP] and a cargo receptor (NCOA4) that is crucial for release of ferritin-bound iron. We also discuss emerging roles of iron in apoptosis and a novel iron-dependent cell death pathway termed ‘ferroptosis’, the dysregulation of iron metabolism in human pathologies, and the use of iron chelators in cancer therapy. Iron is necessary for life, but can also cause cell death. Accordingly, cells evolved a robust, tightly regulated suite of genes for maintaining iron homeostasis. Previous mechanistic studies on iron homeostasis have granted insight into the role of iron in human health and disease. We highlight new regulators of iron metabolism, including iron-trafficking proteins [solute carrier family 39, SLC39, also known as ZRT/IRT-like protein, ZIP; and poly-(rC)-binding protein, PCBP] and a cargo receptor (NCOA4) that is crucial for release of ferritin-bound iron. We also discuss emerging roles of iron in apoptosis and a novel iron-dependent cell death pathway termed ‘ferroptosis’, the dysregulation of iron metabolism in human pathologies, and the use of iron chelators in cancer therapy. Dysregulation of iron metabolism contributes to various human pathologies, including iron overload diseases and cancer.Several new proteins have been identified as crucial iron traffickers and chaperones with important connections to human health.Ferroptosis is a unique cell death pathway that is iron-dependent, non-apoptotic, non-necroptotic, and non-autophagic.Dysregulation of the ferroportin/hepcidin regulatory axis contributes to tumor progression and is predictive of patient outcomes.New iron chelators utilize more specific mechanisms to elicit anticancer activity. Dysregulation of iron metabolism contributes to various human pathologies, including iron overload diseases and cancer. Several new proteins have been identified as crucial iron traffickers and chaperones with important connections to human health. Ferroptosis is a unique cell death pathway that is iron-dependent, non-apoptotic, non-necroptotic, and non-autophagic. Dysregulation of the ferroportin/hepcidin regulatory axis contributes to tumor progression and is predictive of patient outcomes. New iron chelators utilize more specific mechanisms to elicit anticancer activity. Iron is required in a variety of important biological processes including oxygen transport (as heme in hemoglobin), DNA biosynthesis (as a cofactor of ribonucleotide reductase), and ATP generation (as a cofactor for many proteins in the citric acid cycle and electron transport chain); therefore, cells must maintain a sufficient amount of iron. However, iron is redox-active and can generate reactive oxygen species (ROS), leading to oxidative stress and initiation of signaling pathways crucial for cell survival and cell death [1Ray P.D. et al.Reactive oxygen species (ROS) homeostasis and redox regulation in cellular signaling.Cell. Signal. 2012; 24: 981-990Crossref PubMed Scopus (2799) Google Scholar]. To maintain adequate and safe amounts of iron, cells require the coordination of a wide variety of genes, which tightly control both intracellular (reviewed in [2MacKenzie E.L. et al.Intracellular iron transport and storage: from molecular mechanisms to health implications.Antioxid. Redox Signal. 2008; 10: 997-1030Crossref PubMed Scopus (343) Google Scholar, 3Hentze M.W. et al.Two to tango: regulation of mammalian iron metabolism.Cell. 2010; 142: 24-38Abstract Full Text Full Text PDF PubMed Scopus (1407) Google Scholar]) and systemic (reviewed in [4Ganz T. Nemeth E. Hepcidin and iron homeostasis.Biochim. Biophys. Acta. 2012; 1823: 1434-1443Crossref PubMed Scopus (856) Google Scholar]) iron metabolism. Extensive research by many groups has revealed key mechanisms in iron homeostasis (Box 1), as well as links between aberrations in iron homeostasis and human disease. The study of iron metabolism continues to be a dynamic field, with many breakthroughs and novel insights in the past several years. In this review we discuss recent advances in the function and regulation of key iron metabolism genes, including: ferritin (FTH1 and FTL), a protein complex that safely concentrates intracellular iron in a mineralized, redox-inactive form for later use; transferrin (TF), an iron-binding serum protein; transferrin receptor 1 (TfR1, TFRC), a plasma membrane protein that allows cellular uptake of iron-loaded transferrin; divalent metal transporter 1 (DMT1, SLC11A2), a metal transporter that is important for TfR1-mediated iron uptake and dietary iron absorption; ferroportin (Fpn, SLC40A1), the only known cellular iron efflux pump; and hepcidin (HAMP), a circulating peptide hormone that regulates serum iron levels by causing ferroportin degradation. We also examine newly identified regulators in iron metabolism, including new membrane iron transporters and cytosolic iron trafficking proteins. In addition, we review new roles for iron in cell death pathways, as well as the importance of aberrant iron metabolism in human diseases such as cancer.Box 1Control of Mammalian Iron Metabolism Occurs through Two Distinct, but Connected, Regulatory SystemsIntracellular iron metabolism is primarily controlled through coordinated post-transcriptional regulation of various iron metabolism genes (reviewed in [2MacKenzie E.L. et al.Intracellular iron transport and storage: from molecular mechanisms to health implications.Antioxid. Redox Signal. 2008; 10: 997-1030Crossref PubMed Scopus (343) Google Scholar, 3Hentze M.W. et al.Two to tango: regulation of mammalian iron metabolism.Cell. 2010; 142: 24-38Abstract Full Text Full Text PDF PubMed Scopus (1407) Google Scholar]). Many mRNAs involved in iron metabolism contain iron-responsive elements (IREs): stem-loop structures located in 5′- or 3′-untranslated regions (UTRs) flanking the coding sequence (CDS) [76Kuhn L.C. Iron regulatory proteins and their role in controlling iron metabolism.Metallomics: Int. Bio. Sci. 2015; 7: 232-243Crossref PubMed Google Scholar]. IREs bind two functionally similar iron regulatory proteins, IRP1 and IRP2. Depending upon whether the IRE is located within the 5′-UTR or in the 3′-UTR, the IRE–IRP interaction has opposite effects on target gene expression. In low iron conditions, 5′-UTR IREs are translationally repressed as a result of IRP blocking ribosome recruitment, whereas 3′-UTR IREs mediate enhanced mRNA stability, ultimately increasing protein levels (Figure IA, Low iron). High iron removes the IRPs, facilitating translational activation of 5′-UTR IRE mRNAs or degradation of 3′-UTR IRE mRNAs via endonuclease attack (Figure IA, High iron). 5′-UTR IREs are usually found in genes that lower the amount of cellular labile iron (i.e., iron that is unbound and redox active) such as ferritin and ferroportin, whereas 3′-UTR IREs are found in genes that facilitate iron uptake such as transferrin receptor 1 and divalent metal transporter 1 (Figure IA) Classically, it was believed that iron affected the IRE–IRP interaction entirely through effects on the IRPs. However, the ability of Fe2+ (and other metal ions) to directly bind to the IRE stem-loop was recently observed [77Khan M.A. et al.Direct Fe2+ sensing by iron-responsive messenger RNA:repressor complexes weakens binding.J. Biol. Chem. 2009; 284: 30122-30128Crossref PubMed Scopus (34) Google Scholar]. Fe2+ binding to 5′-UTR IREs induces a conformational change in the stem-loop, which decreases its affinity for IRPs [78Ma J. et al.Fe2+ binds iron responsive element-RNA, selectively changing protein-binding affinities and regulating mRNA repression and activation.Proc. Natl. Acad. Sci. U.S.A. 2012; 109: 8417-8422Crossref PubMed Scopus (50) Google Scholar] and increases its affinity for a ribosome recruitment factor eIF4F [79Khan M.A. et al.Rapid kinetics of iron responsive element (IRE) RNA/iron regulatory protein 1 and IRE-RNA/eIF4F complexes respond differently to metal ions.Nucleic Acids Res. 2014; 42: 6567-6577Crossref PubMed Scopus (18) Google Scholar], showing that the IRE structure itself contributes to post-transcriptional control of gene expression.In mammals, systemic iron homeostasis is controlled by the hepatocyte-secreted hormone hepcidin (reviewed in [4Ganz T. Nemeth E. Hepcidin and iron homeostasis.Biochim. Biophys. Acta. 2012; 1823: 1434-1443Crossref PubMed Scopus (856) Google Scholar]). Hepcidin circulates in the serum and binds to the cellular iron exporter ferroportin (Fpn), stimulating Fpn degradation and leading to cellular retention of iron (Figure IB). Hepcidin expression is directly correlated with both cellular and serum iron statuses and is controlled through a complex iron-sensing signaling pathway (reviewed in [4Ganz T. Nemeth E. Hepcidin and iron homeostasis.Biochim. Biophys. Acta. 2012; 1823: 1434-1443Crossref PubMed Scopus (856) Google Scholar]). Elevated serum hepcidin downregulates Fpn in duodenal enterocytes (which are responsible for dietary iron absorption), macrophages (which contain large amounts of iron from erythrocyte recycling), and hepatocytes (which act as an iron reservoir and export iron as needed). This leads to an overall reduction in serum iron (Figure IB). Some overlap exists between the IRE–IRP system and the hepcidin–Fpn axis (reviewed in [3Hentze M.W. et al.Two to tango: regulation of mammalian iron metabolism.Cell. 2010; 142: 24-38Abstract Full Text Full Text PDF PubMed Scopus (1407) Google Scholar]) because Fpn is regulated by IRPs though its 5′ -UTR IRE. Intracellular iron metabolism is primarily controlled through coordinated post-transcriptional regulation of various iron metabolism genes (reviewed in [2MacKenzie E.L. et al.Intracellular iron transport and storage: from molecular mechanisms to health implications.Antioxid. Redox Signal. 2008; 10: 997-1030Crossref PubMed Scopus (343) Google Scholar, 3Hentze M.W. et al.Two to tango: regulation of mammalian iron metabolism.Cell. 2010; 142: 24-38Abstract Full Text Full Text PDF PubMed Scopus (1407) Google Scholar]). Many mRNAs involved in iron metabolism contain iron-responsive elements (IREs): stem-loop structures located in 5′- or 3′-untranslated regions (UTRs) flanking the coding sequence (CDS) [76Kuhn L.C. Iron regulatory proteins and their role in controlling iron metabolism.Metallomics: Int. Bio. Sci. 2015; 7: 232-243Crossref PubMed Google Scholar]. IREs bind two functionally similar iron regulatory proteins, IRP1 and IRP2. Depending upon whether the IRE is located within the 5′-UTR or in the 3′-UTR, the IRE–IRP interaction has opposite effects on target gene expression. In low iron conditions, 5′-UTR IREs are translationally repressed as a result of IRP blocking ribosome recruitment, whereas 3′-UTR IREs mediate enhanced mRNA stability, ultimately increasing protein levels (Figure IA, Low iron). High iron removes the IRPs, facilitating translational activation of 5′-UTR IRE mRNAs or degradation of 3′-UTR IRE mRNAs via endonuclease attack (Figure IA, High iron). 5′-UTR IREs are usually found in genes that lower the amount of cellular labile iron (i.e., iron that is unbound and redox active) such as ferritin and ferroportin, whereas 3′-UTR IREs are found in genes that facilitate iron uptake such as transferrin receptor 1 and divalent metal transporter 1 (Figure IA) Classically, it was believed that iron affected the IRE–IRP interaction entirely through effects on the IRPs. However, the ability of Fe2+ (and other metal ions) to directly bind to the IRE stem-loop was recently observed [77Khan M.A. et al.Direct Fe2+ sensing by iron-responsive messenger RNA:repressor complexes weakens binding.J. Biol. Chem. 2009; 284: 30122-30128Crossref PubMed Scopus (34) Google Scholar]. Fe2+ binding to 5′-UTR IREs induces a conformational change in the stem-loop, which decreases its affinity for IRPs [78Ma J. et al.Fe2+ binds iron responsive element-RNA, selectively changing protein-binding affinities and regulating mRNA repression and activation.Proc. Natl. Acad. Sci. U.S.A. 2012; 109: 8417-8422Crossref PubMed Scopus (50) Google Scholar] and increases its affinity for a ribosome recruitment factor eIF4F [79Khan M.A. et al.Rapid kinetics of iron responsive element (IRE) RNA/iron regulatory protein 1 and IRE-RNA/eIF4F complexes respond differently to metal ions.Nucleic Acids Res. 2014; 42: 6567-6577Crossref PubMed Scopus (18) Google Scholar], showing that the IRE structure itself contributes to post-transcriptional control of gene expression. In mammals, systemic iron homeostasis is controlled by the hepatocyte-secreted hormone hepcidin (reviewed in [4Ganz T. Nemeth E. Hepcidin and iron homeostasis.Biochim. Biophys. Acta. 2012; 1823: 1434-1443Crossref PubMed Scopus (856) Google Scholar]). Hepcidin circulates in the serum and binds to the cellular iron exporter ferroportin (Fpn), stimulating Fpn degradation and leading to cellular retention of iron (Figure IB). Hepcidin expression is directly correlated with both cellular and serum iron statuses and is controlled through a complex iron-sensing signaling pathway (reviewed in [4Ganz T. Nemeth E. Hepcidin and iron homeostasis.Biochim. Biophys. Acta. 2012; 1823: 1434-1443Crossref PubMed Scopus (856) Google Scholar]). Elevated serum hepcidin downregulates Fpn in duodenal enterocytes (which are responsible for dietary iron absorption), macrophages (which contain large amounts of iron from erythrocyte recycling), and hepatocytes (which act as an iron reservoir and export iron as needed). This leads to an overall reduction in serum iron (Figure IB). Some overlap exists between the IRE–IRP system and the hepcidin–Fpn axis (reviewed in [3Hentze M.W. et al.Two to tango: regulation of mammalian iron metabolism.Cell. 2010; 142: 24-38Abstract Full Text Full Text PDF PubMed Scopus (1407) Google Scholar]) because Fpn is regulated by IRPs though its 5′ -UTR IRE. In recent years several new proteins have been identified as key players in intracellular iron trafficking and utilization. This section discusses novel iron transporters, iron chaperones, and ferritin-shuttling proteins. The solute carrier family 39 (SLC39), also known as the ZRT/IRT-like protein (ZIP) family, contains 14 members and has classically been understood to comprise transmembrane zinc transporters that pump extracellular Zn2+ into the cell [5Jeong J. Eide D.J. The SLC39 family of zinc transporters.Mol. Aspects Med. 2013; 34: 612-619Crossref PubMed Scopus (279) Google Scholar]. However, recent studies have uncovered a role for two ZIP family proteins in iron transport [6Jenkitkasemwong S. et al.Physiologic implications of metal-ion transport by ZIP14 and ZIP8.Biometals. 2012; 25: 643-655Crossref PubMed Scopus (169) Google Scholar]. ZIP14 was found to mediate the uptake of non-transferrin-bound iron (NTBI) [7Liuzzi J.P. et al.Zip14 (Slc39a14) mediates non-transferrin-bound iron uptake into cells.Proc. Natl. Acad. Sci. U.S.A. 2006; 103: 13612-13617Crossref PubMed Scopus (411) Google Scholar] by directly transporting NTBI across the cell membrane [8Pinilla-Tenas J.J. et al.Zip14 is a complex broad-scope metal-ion transporter whose functional properties support roles in the cellular uptake of zinc and nontransferrin-bound iron.Am J. Physiol. Cell Physiol. 2011; 301: C862-C871Crossref PubMed Scopus (156) Google Scholar] (Figure 1, left, ‘Uptake’). ZIP14-mediated NTBI transport is enhanced by the Fe3+ to Fe2+ ferrireductase activity of the prion protein PRNP [9Tripathi A.K. et al.Prion protein functions as a ferrireductase partner for ZIP14 and DMT1.Free Radic. Biol. Med. 2015; 84: 322-330Crossref PubMed Scopus (48) Google Scholar]. ZIP14 is also capable of transporting transferrin-bound iron from the endosome to the cytoplasm, similarly to DMT1 [10Zhao N. et al.ZRT/IRT-like protein 14 (ZIP14) promotes the cellular assimilation of iron from transferrin.J. Biol. Chem. 2010; 285: 32141-32150Crossref PubMed Scopus (126) Google Scholar], although subsequent studies have revealed that this is likely not the major function of ZIP14 [11Ji C. Kosman D.J. Molecular mechanisms of non-transferrin-bound and transferring-bound iron uptake in primary hippocampal neurons.J. Neurochem. 2015; 133: 668-683Crossref PubMed Scopus (57) Google Scholar]. ZIP8, the closest relative to ZIP14 [6Jenkitkasemwong S. et al.Physiologic implications of metal-ion transport by ZIP14 and ZIP8.Biometals. 2012; 25: 643-655Crossref PubMed Scopus (169) Google Scholar], was also found to transport NTBI [12Wang C.Y. et al.ZIP8 is an iron and zinc transporter whose cell-surface expression is up-regulated by cellular iron loading.J. Biol. Chem. 2012; 287: 34032-34043Crossref PubMed Scopus (254) Google Scholar] (Figure 1, left, ‘Uptake’). Interestingly, both ZIP8 and ZIP14 show maximal iron transport above pH 7, near physiological pH [8Pinilla-Tenas J.J. et al.Zip14 is a complex broad-scope metal-ion transporter whose functional properties support roles in the cellular uptake of zinc and nontransferrin-bound iron.Am J. Physiol. Cell Physiol. 2011; 301: C862-C871Crossref PubMed Scopus (156) Google Scholar, 12Wang C.Y. et al.ZIP8 is an iron and zinc transporter whose cell-surface expression is up-regulated by cellular iron loading.J. Biol. Chem. 2012; 287: 34032-34043Crossref PubMed Scopus (254) Google Scholar], whereas DMT1 is most efficient at pH 5.5 [13Gunshin H. et al.Cloning and characterization of a mammalian proton-coupled metal-ion transporter.Nature. 1997; 388: 482-488Crossref PubMed Scopus (2652) Google Scholar], which corresponds to the pH of acidified endosome. The widely disparate pH activities suggest that ZIP8/14 and DMT1 may have different biological roles. Indeed, a recent report utilizing primary rat hippocampal neurons showed that ZIP8 is the major NTBI transporter whereas DMT1 was principally responsible for transferrin-bound iron transport from the endosome [11Ji C. Kosman D.J. Molecular mechanisms of non-transferrin-bound and transferring-bound iron uptake in primary hippocampal neurons.J. Neurochem. 2015; 133: 668-683Crossref PubMed Scopus (57) Google Scholar]. ZIP8 and ZIP14 appear to have nonredundant functions because they have distinct tissue expression profiles and knockout mouse models show different phenotypes. Knockout of ZIP8 in mice causes failures in hematopoiesis and organogenesis of the spleen, liver, kidney, and lung, with associated perinatal lethality [14Galvez-Peralta M. et al.ZIP8 zinc transporter: indispensable role for both multiple-organ organogenesis and hematopoiesis in utero.PLoS ONE. 2012; 7: e36055Crossref PubMed Scopus (81) Google Scholar]. Zip8−/− mice exhibited profound anemia and have significantly lowered amounts of iron and zinc in various tissues, with the hematopoietic defect seemingly being caused by the combination of iron and zinc deficiency. By contrast, Zip14−/− mice are viable and have adequate iron stores, but exhibit dwarfism owing to aberrant bone growth. The dwarfism was found to primarily be the result of altered zinc, rather than iron, metabolism, indicating that zinc transport is more important than iron transport by ZIP14 for normal growth [15Hojyo S. et al.The zinc transporter SLC39A14/ZIP14 controls G-protein coupled receptor-mediated signaling required for systemic growth.PLoS ONE. 2011; 6: e18059Crossref PubMed Scopus (130) Google Scholar]. However, a more recent study uncovered a crucial role for ZIP14 in iron transport by knocking out ZIP14 in two mouse models for hereditary hemochromatosis, a family of iron overload diseases [16Jenkitkasemwong S. et al.SLC39A14 is required for the development of hepatocellular iron overload in murine models of hereditary hemochromatosis.Cell Metab. 2015; 22: 138-150Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar]. Hemochromatosis results in a progressive accumulation of iron primarily in the liver, resulting in hepatic fibrosis and potentially hepatocellular carcinoma. Knocking out ZIP14 in Hfe−/− (hereditary hemochromatosis protein) or Hfe2−/− (hemojuvelin) mice [modeling type 1 and type 2 (juvenile) hemochromatosis, respectively] prevented iron accumulation in the liver. ZIP14 knockout also prevented iron overload in Hfe2−/− mouse pancreas, but not heart [16Jenkitkasemwong S. et al.SLC39A14 is required for the development of hepatocellular iron overload in murine models of hereditary hemochromatosis.Cell Metab. 2015; 22: 138-150Abstract Full Text Full Text PDF PubMed Scopus (146) Google Scholar], which could be attributable to the high pancreatic but low cardiac expression of ZIP14 [17Nam H. et al.ZIP14 and DMT1 in the liver, pancreas, and heart are differentially regulated by iron deficiency and overload: implications for tissue iron uptake in iron-related disorders.Haematologica. 2013; 98: 1049-1057Crossref PubMed Scopus (121) Google Scholar]. Uncovering the importance of ZIP14 in iron loading provides a new target in the treatment of iron overload diseases. Namely, inhibition of ZIP14 could potentially block hepatic and pancreatic iron overloading, and prevent subsequent organ damage. The expression of ZIP8 and ZIP14 is regulated differently than other iron transporter genes such as TfR1 and DMT1, both of which are controlled post-transcriptionally by changes of mRNA stability through the 3′ untranslated region (3′ -UTR) iron-responsive element (IRE)–iron regulatory protein (IRP) interaction (Box 1) [2MacKenzie E.L. et al.Intracellular iron transport and storage: from molecular mechanisms to health implications.Antioxid. Redox Signal. 2008; 10: 997-1030Crossref PubMed Scopus (343) Google Scholar, 3Hentze M.W. et al.Two to tango: regulation of mammalian iron metabolism.Cell. 2010; 142: 24-38Abstract Full Text Full Text PDF PubMed Scopus (1407) Google Scholar]. By contrast, ZIP8 and ZIP14 mRNA stability is unaffected by iron, and ZIP protein levels are positively correlated with iron status, in that iron loading increases ZIP8/14 protein amount whereas iron deficiency decreases protein amount [12Wang C.Y. et al.ZIP8 is an iron and zinc transporter whose cell-surface expression is up-regulated by cellular iron loading.J. Biol. Chem. 2012; 287: 34032-34043Crossref PubMed Scopus (254) Google Scholar, 18Zhao N. et al.An iron-regulated and glycosylation-dependent proteasomal degradation pathway for the plasma membrane metal transporter ZIP14.Proc. Natl. Acad. Sci. U.S.A. 2014; 111: 9175-9180Crossref PubMed Scopus (47) Google Scholar]. In summary, ZIP8 and ZIP14 are newly recognized iron transporters with important roles in the uptake of NTBI and in iron overload diseases. Further investigation will be necessary to determine the importance of ZIP iron transport in other contexts, such as disease states with disordered iron homeostasis (anemia, cancer, etc.). PCBPs are a family of four proteins that were originally identified for their high affinity binding to tracts of polycytosine and have been studied as regulators of gene expression (reviewed in [19Chaudhury A. et al.Heterogeneous nuclear ribonucleoproteins (hnRNPs) in cellular processes: focus on hnRNP E1's multifunctional regulatory roles.RNA. 2010; 16: 1449-1462Crossref PubMed Scopus (197) Google Scholar]). Each member of the PCBP family contains three highly conserved K homology (KH) domains and can form complexes with other family members. Recently, PCBP1 and PCBP2 were identified as cytosolic iron chaperones that are responsible for delivering iron to ferritin, the protein that is primarily responsible for storing intracellular iron in a non-toxic, mineralized state [20Shi H. et al.A cytosolic iron chaperone that delivers iron to ferritin.Science. 2008; 320: 1207-1210Crossref PubMed Scopus (365) Google Scholar, 21Leidgens S. et al.Each member of the poly-r(C)-binding protein 1 (PCBP) family exhibits iron chaperone activity toward ferritin.J. Biol. Chem. 2013; 288: 17791-17802Crossref PubMed Scopus (128) Google Scholar] (Figure 1, center, ‘Distribution’). PCBP3 and PCBP4 are expressed at much lower levels, with a limited tissue distribution and an unclear biological role, but there is some evidence that they can also act as iron chaperones and that PCBP3 can bind to ferritin [21Leidgens S. et al.Each member of the poly-r(C)-binding protein 1 (PCBP) family exhibits iron chaperone activity toward ferritin.J. Biol. Chem. 2013; 288: 17791-17802Crossref PubMed Scopus (128) Google Scholar]. To date, the majority of the research on the iron chaperone function of the PCBP family has focused on PCBP1 and PCBP2. Both PCBP1 and PCBP2 are necessary to form a stable ternary complex with ferritin, and ferritin–PCBP interactions were dependent upon iron-loaded PCBP1/2 [21Leidgens S. et al.Each member of the poly-r(C)-binding protein 1 (PCBP) family exhibits iron chaperone activity toward ferritin.J. Biol. Chem. 2013; 288: 17791-17802Crossref PubMed Scopus (128) Google Scholar]. In addition to ferritin, PCBP1 and PCBP2 can also deliver iron to proteins that require non-heme iron as a cofactor (‘metallation’, Figure 1, center, ‘Distribution’), such as hypoxia-inducible factor (HIF) prolyl and asparginyl hydoxylases [22Nandal A. et al.Activation of the HIF prolyl hydroxylase by the iron chaperones PCBP1 and PCBP2.Cell Metab. 2011; 14: 647-657Abstract Full Text Full Text PDF PubMed Scopus (150) Google Scholar], and deoxyhypusine hydroxylase [23Frey A.G. et al.Iron chaperones PCBP1 and PCBP2 mediate the metallation of the dinuclear iron enzyme deoxyhypusine hydroxylase.Proc. Natl. Acad. Sci. U.S.A. 2014; 111: 8031-8036Crossref PubMed Scopus (86) Google Scholar]. PCBP2 is loaded with iron through binding to the iron transporter DMT1. Iron transfer to PCBP2 is mediated through interaction of the second KH domain of PCBP2 with the N-terminal cytoplasmic domain of DMT1 [24Yanatori I. et al.Chaperone protein involved in transmembrane transport of iron.Biochem. J. 2014; 462: 25-37Crossref PubMed Scopus (88) Google Scholar]. After transfer of Fe2+, PCBP2 subsequently dissociates from DMT1 (Figure 1, center, ‘Distribution’). By contrast, PCPB1 does not interact with DMT1, and the mechanism of apo-PCBP1 iron acquisition is still undetermined. A comprehensive screen of both iron-loaded and apo-PCBP1 and -2 binding to other iron metabolism genes (ZIP transporters, etc.) is crucial for understanding overall cytoplasmic iron shuttling and may shed light on PCBP1 iron loading. Many questions about the roles and mechanisms of the PCBP proteins in iron homeostasis remain unanswered. For instance, it was noted that PCBP2 can also bind to the iron exporter ferroportin (Fpn) [24Yanatori I. et al.Chaperone protein involved in transmembrane transport of iron.Biochem. J. 2014; 462: 25-37Crossref PubMed Scopus (88) Google Scholar] (Figure 1, center, ‘Distribution’), but that association remains to be characterized in depth, including what effects, if any, PCBP2 has on Fpn iron export. In addition, the question of what roles, if any, PCBP3 and PCBP4 have in intracellular iron homeostasis remains to be answered. Ferritin stores ferric iron in a mineralized, non-toxic state. However, ferritin-bound iron cannot be utilized by the cell. Consequently, iron must be released from ferritin to be biologically useful, usually through lysosomal degradation of ferritin [25Kidane T.Z. et al.Release of iron from ferritin requires lysosomal activity.Am J. Physiol. Cell Physiol. 2006; 291: C445-C455Crossref PubMed Scopus (187) Google Scholar, 26Asano T. et al.Distinct mechanisms of ferritin delivery to lysosomes in iron-depleted and iron-replete cells.Mol. Cell. 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While the rate of ferritin degradation appears to be the same in iron-deficient and iron-replete conditions, the delivery mechanism of ferritin to lysosomes seems to be different: autophagy is responsible for delivering ferritin to the lysosome during iron deficiency, whereas a non-autophagic pathway dominates during iron sufficiency [26Asano T. et al.Distinct mechanisms of ferritin delivery to lysosomes in iron-depleted and iron-replete cells.Mol. Cell. Biol. 2011; 31: 2040-2052Crossref PubMed Scopus (164) Google Scholar]. Recently, nuclear receptor coactivator 4 (NCOA4) was identified as the protein responsible for mediating ferritin autophagy [29Dowdle W.E. et al.Selective VPS34 inhibitor blocks autophagy and uncovers a role for NCOA4 in ferritin degradation and iron homeostasis in vivo.Nat. Cell Biol. 2014; 16: 1069-1079Crossref PubMed Scopus (415) Google Scholar, 30Mancias J.D. et al.Quantitative proteomics identifies NCOA4 as the cargo receptor mediating ferritinophagy.Nature. 2014; 509: 105-109Crossref PubMed Scopus (809) Google Scholar]. NCOA4 was originally identified as a coactivator of several nuclear receptors but was later found to primarily be localized in the cytoplasm and have roles in various biological processes independent of its coactivator function (reviewed in [31Kollara A. Brown T.J. Expression and function of nuclear receptor co-activato