Nuclear translocation of STAT5 initiates iron overload in huntington's disease by up-regulating IRP1 expression.

Mutant huntingtin (mHtt) proteins interact to form aggregates, disrupting cellular functions including transcriptional dysregulation and iron imbalance in patients with Huntington's disease (HD) and mouse disease models. Previous studies have indicated that mHtt may lead to abnormal iron homeostasis by upregulating the expression of iron response protein 1 (IRP1) in the striatum and cortex of N171-82Q HD transgenic mice, as well as in HEK293 cells expressing the N-terminal fragment of mHtt containing 160 CAG repeats. However, the mechanism underlying the upregulation of IRP1 remains unclear. We investigated the levels and phosphorylation status of signal transducer and activator of transcription 5 (STAT5) in the brains of N171-82Q HD transgenic mice using immunohistochemistry staining. We also assessed the nuclear localization of STAT5 protein through western blot and immunofluorescence, and measured the relative RNA expression levels of STAT5 and IRP1 using RT-PCR in both N171-82Q HD transgenic mice and HEK293 cells expressing the N-terminal fragment of huntingtin. Our findings demonstrate that the transcription factor STAT5 regulates the transcription of the IPR1 gene in HEK293 cells. Notably, both the brains of N171-82Q mice and 160Q HEK293 cells exhibited increased nuclear content of STAT5, despite unchanged total STAT5 expression. These results suggest that mHtt promotes the nuclear translocation of STAT5, leading to enhanced expression of IRP1. The nuclear translocation of STAT5 initiates abnormal iron homeostatic pathways, characterized by elevated IRP1 expression, increased levels of transferrin and transferrin receptor, and iron accumulation in the brains of HD mice. These findings provide valuable insights into potential therapeutic strategies targeting iron homeostasis in HD.

Using Biotinylated Iron-Responsive Element to Analyze the Activity of Iron Regulatory Proteins.

Iron regulatory proteins (IRP1 and IRP2) are the master regulators of mammalian iron homeostasis. They bind to the iron-responsive elements (IREs) of the transcripts of iron-related genes to regulate their expression, thereby maintaining cellular iron availability. The primary method to measure the IRE-binding activity of IRPs is the electrophoresis mobility shift assay (EMSA). This method is particularly useful for evaluating IRP1 activity, since IRP1 is a bifunctional enzyme and its protein levels remain similar during conversion between the IRE-binding protein and cytosolic aconitase forms. Here, we exploited a method of using a biotinylated-IRE probe to separate IRE-binding IRPs followed by immunoblotting to analyze the IRE-binding activity. This method allows for the successful measurement of IRP activity in cultured cells and mouse tissues under various iron conditions. By separating IRE-binding IRPs from the rest of the lysates, this method increases the specificity of IRP antibodies and verifies whether a band represents an IRP, thereby revealing some previously unrecognized information about IRPs. With this method, we showed that the S711-phosphorylated IRP1 was found only in the IRE-binding form in PMA-treated Hep3B cells. Second, we found a truncated IRE-binding IRP2 isoform that is generated by proteolytic cleavage on sites in the 73aa insert region of the IRP2 protein. Third, we found that higher levels of SDS, compared to 1-2% SDS in regular loading buffer, could dramatically increase the band intensity of IRPs in immunoblots, especially in HL-60 cells. Fourth, we found that the addition of SDS or LDS to cell lysates activated protein degradation at 37 °C or room temperature, especially in HL-60 cell lysates. As this method is more practical, sensitive, and cost-effective, we believe that its application will enhance future research on iron regulation and metabolism.

CD63 is regulated by iron via the IRE-IRP system and is important for ferritin secretion by extracellular vesicles.

Extracellular vesicles (EVs) transfer functional molecules between cells. CD63 is a widely recognized EV marker that contributes to EV secretion from cells. However, the regulation of its expression remains largely unknown. Ferritin is a cellular iron storage protein that can also be secreted by the exosome pathway, and serum ferritin levels classically reflect body iron stores. Iron metabolism-associated proteins such as ferritin are intricately regulated by cellular iron levels via the iron responsive element-iron regulatory protein (IRE-IRP) system. Herein, we present a novel mechanism demonstrating that the expression of the EV-associated protein CD63 is under the regulation of the IRE-IRP system. We discovered a canonical IRE in the 5' untranslated region of CD63 messenger RNA that is responsible for regulating its expression in response to increased iron. Cellular iron loading caused a marked increase in CD63 expression and the secretion of CD63+ EVs from cells, which were shown to contain ferritin-H and ferritin-L. Our results demonstrate that under iron loading, intracellular ferritin is transferred via nuclear receptor coactivator 4 (NCOA4) to CD63+ EVs that are then secreted. Such iron-regulated secretion of the major iron storage protein ferritin via CD63+ EVs, is significant for understanding the local cell-to-cell exchange of ferritin and iron.

The thermogenic characteristics of adipocytes are dependent on the regulation of iron homeostasis.

The development of thermogenic adipocytes concurs with mitochondrial biogenesis, an iron-dependent pathway. Iron regulatory proteins (IRP) 1 and 2 are RNA-binding proteins that regulate intracellular iron homeostasis. IRPs bind to the iron-response element (IRE) of their target mRNAs, balancing iron uptake and deposition at the posttranscriptional levels. However, IRP/IRE-dependent iron regulation in adipocytes is largely unknown. We hypothesized that iron demands are higher in brown/beige adipocytes than white adipocytes to maintain the thermogenic mitochondrial capacity. To test this hypothesis, we investigated the IRP/IRE regulatory system in different depots of adipose tissue. Our results revealed that 1) IRP/IRE interaction was increased in proportional to the thermogenic function of the adipose depot, 2) adipose iron content was increased in adipose tissue browning upon ß3-adrenoceptor stimulation, while decreased in thermoneutral conditions, and 3) modulation of iron content was linked with mitochondrial biogenesis. Moreover, the iron requirement was higher in HIB1B brown adipocytes than 3T3-L1 white adipocytes during differentiation. The reduction of the labile iron pool (LIP) suppressed the differentiation of brown/beige adipocytes and mitochondrial biogenesis. Using the 59Fe-Tf, we also demonstrated that thermogenic stimuli triggered cell-autonomous iron uptake and mitochondrial compartmentalization as well as enhanced mitochondrial respiration. Collectively, our work demonstrated that IRP/IRE signaling and subsequent adaptation in iron metabolism are a critical determinant for the thermogenic function of adipocytes.

Dysregulation of the sensory and regulatory pathways controlling cellular iron metabolism in unilateral obstructive nephropathy.

Chronic kidney disease involves disturbances in iron metabolism including anemia caused by insufficient erythropoietin (EPO) production. However, underlying mechanisms responsible for the dysregulation of cellular iron metabolism are incompletely defined. Using the unilateral ureteral obstruction (UUO) model in Irp1+/+ and Irp1-/- mice, we asked if iron regulatory proteins (IRPs), the central regulators of cellular iron metabolism and suppressors of EPO production, contribute to the etiology of anemia in kidney failure. We identified a significant reduction in IRP protein level and RNA binding activity that associates with a loss of the iron uptake protein transferrin receptor 1 (TfR1), increased expression of the iron storage protein subunits H- and L-ferritin, and a low but overall variable level of stainable iron in the obstructed kidney. This reduction in IRP RNA binding activity and ferritin RNA levels suggests the concomitant rise in ferritin expression and iron content in kidney failure is IRP dependent. In contrast, the reduction in the Epo mRNA level in the obstructed kidney was not rescued by genetic ablation of IRP1, suggesting disruption of normal hypoxia-inducible factor (HIF)-2α regulation. Furthermore, reduced expression of some HIF-α target genes in UUO occurred in the face of increased expression of HIF-α proteins and prolyl hydroxylases 2 and 1, the latter of which is not known to be HIF-α mediated. Our results suggest that the IRP system drives changes in cellular iron metabolism that are associated with kidney failure in UUO but that the impact of IRPs on EPO production is overridden by disrupted hypoxia signaling.NEW & NOTEWORTHY This study demonstrates that iron metabolism and hypoxia signaling are dysregulated in unilateral obstructive nephropathy. Expression of iron regulatory proteins (IRPs), central regulators of cellular iron metabolism, and the iron uptake (transferrin receptor 1) and storage (ferritins) proteins they target is strongly altered. This suggests a role of IRPs in previously observed changes in iron metabolism in progressive renal disease. Hypoxia signaling is disrupted and appeared to dominate the action of IRP1 in controlling erythropoietin expression.

Function and crystal structure of the dimeric P-loop ATPase CFD1 coordinating an exposed [4Fe-4S] cluster for transfer to apoproteins.

Maturation of iron-sulfur (Fe-S) proteins in eukaryotes requires complex machineries in mitochondria and cytosol. Initially, Fe-S clusters are assembled on dedicated scaffold proteins and then are trafficked to target apoproteins. Within the cytosolic Fe-S protein assembly (CIA) machinery, the conserved P-loop nucleoside triphosphatase Nbp35 performs a scaffold function. In yeast, Nbp35 cooperates with the related Cfd1, which is evolutionary less conserved and is absent in plants. Here, we investigated the potential scaffold function of human CFD1 (NUBP2) in CFD1-depleted HeLa cells by measuring Fe-S enzyme activities or 55Fe incorporation into Fe-S target proteins. We show that CFD1, in complex with NBP35 (NUBP1), performs a crucial role in the maturation of all tested cytosolic and nuclear Fe-S proteins, including essential ones involved in protein translation and DNA maintenance. CFD1 also matures iron regulatory protein 1 and thus is critical for cellular iron homeostasis. To better understand the scaffold function of CFD1-NBP35, we resolved the crystal structure of Chaetomium thermophilum holo-Cfd1 (ctCfd1) at 2.6-Å resolution as a model Cfd1 protein. Importantly, two ctCfd1 monomers coordinate a bridging [4Fe-4S] cluster via two conserved cysteine residues. The surface-exposed topology of the cluster is ideally suited for both de novo assembly and facile transfer to Fe-S apoproteins mediated by other CIA factors. ctCfd1 specifically interacted with ATP, which presumably associates with a pocket near the Cfd1 dimer interface formed by the conserved Walker motif. In contrast, ctNbp35 preferentially bound GTP, implying differential regulation of the two fungal scaffold components during Fe-S cluster assembly and/or release.

The Role of GSH in Intracellular Iron Trafficking.

Evidence is reviewed for the role of glutathione in providing a ligand for the cytosolic iron pool. The possibility of histidine and carnosine forming ternary complexes with iron(II)glutathione is discussed and the physiological significance of these interactions considered. The role of carnosine in muscle, brain, and kidney physiology is far from established and evidence is presented that the iron(II)-binding capability of carnosine relates to this role.

Exploiting the passenger ACO1-deficiency arising from 9p21 deletions to kill T-cell lymphoblastic neoplasia cells.

Precursor T-cell lymphoblastic neoplasms are aggressive malignancies in need for more effective and specific therapeutic treatments. A significant fraction of these neoplasms harbor deletions on the locus 9p21, targeting the tumor suppressor CDKN2A but also deleting the aconitase 1 (ACO1) gene, a neighboring housekeeping gene involved in cytoplasm and mitochondrial metabolism. Here we show that reducing the aconitase activity with fluorocitrate decreases the viability of T-cell lymphoblastic neoplasia cells in correlation to the differential aconitase expression. The consequences of the treatment were evidenced in vitro using T-cell lymphoblastic neoplasia cell lines exhibiting 9p21 deletions and variable levels of ACO1 expression or activity. Similar results were observed in melanoma cell lines, suggesting a true potential for fluorocitrate in different cancer types. Notably, ectopic expression of ACO1 alleviated the susceptibility of cell lines to fluorocitrate and, conversely, knockdown experiments increased susceptibility of resistant cell lines. These findings were confirmed in vivo on athymic nude mice by using tumor xenografts derived from two T-cell lines with different levels of ACO1. Taken together, our results indicate that the non-targeted ACO1 deficiency induced by common deletions exerts a collateral cellular lethality that can be used as a novel therapeutic strategy in the treatment of several types of cancer.

In vivo bioluminescence imaging of labile iron accumulation in a murine model of Acinetobacter baumannii infection.

Iron is an essential metal for all organisms, yet disruption of its homeostasis, particularly in labile forms that can contribute to oxidative stress, is connected to diseases ranging from infection to cancer to neurodegeneration. Iron deficiency is also among the most common nutritional deficiencies worldwide. To advance studies of iron in healthy and disease states, we now report the synthesis and characterization of iron-caged luciferin-1 (ICL-1), a bioluminescent probe that enables longitudinal monitoring of labile iron pools (LIPs) in living animals. ICL-1 utilizes a bioinspired endoperoxide trigger to release d-aminoluciferin for selective reactivity-based detection of Fe2+ with metal and oxidation state specificity. The probe can detect physiological changes in labile Fe2+ levels in live cells and mice experiencing iron deficiency or overload. Application of ICL-1 in a model of systemic bacterial infection reveals increased iron accumulation in infected tissues that accompany transcriptional changes consistent with elevations in both iron acquisition and retention. The ability to assess iron status in living animals provides a powerful technology for studying the contributions of iron metabolism to physiology and pathology.

Nuclear aconitase antagonizes heterochromatic silencing by interfering with Chp1 binding to DNA.

In Schizosaccharomyces pombe, there are two aconitases, Aco1 and Aco2, involved in the Krebs cycle in mitochondria. Interestingly, Aco2 is localized to nucleus as well. Here, we investigated the nuclear role of Aco2 by deleting its nuclear localization signal. The aco2ΔNLS mutation suppressed the gene-silencing defects of RNAi mutants at the centromere, where heterochromatin formation depends on RNAi pathway. In Δago1, the aco2ΔNLS mutation restored heterochromatin through elevating Chp1 binding. Aco2 physically interacted with Chp1 via the N-terminal chromodomain that binds to methylated histone H3K9. In the sub-telomeric region, where heterochromatin forms independent of RNAi pathway, the single aco2ΔNLS mutation caused extra gene silencing via elevating Chp1 binding, without increasing histone methylation. The anti-silencing effect did not require the catalytic function of aconitase. Taken together, Aco2 functions as an epigenetic regulator of gene expression, through associating with chromodomain of Chp1 to maintain heterochromatin.

Consequences of cytochrome c oxidase assembly defects for the yeast stationary phase.

The assembly of cytochrome c oxidase (COX) is essential for a functional mitochondrial respiratory chain, although the consequences of a loss of assembled COX at yeast stationary phase, an excellent model for terminally differentiated cells in humans, remain largely unexamined. In this study, we show that a wild-type respiratory competent yeast strain at stationary phase is characterized by a decreased oxidative capacity, as seen by a reduction in the amount of assembled COX and by a decrease in protein levels of several COX assembly factors. In contrast, loss of assembled COX results in the decreased abundance of many mitochondrial proteins at stationary phase, which is likely due to decreased membrane potential and changes in mitophagy. In addition to an altered mitochondrial proteome, COX assembly mutants display unexpected changes in markers of cellular oxidative stress at stationary phase. Our results suggest that mitochondria may not be a major source of reactive oxygen species at stationary phase in cells lacking an intact respiratory chain.

The roles of hypoxia-inducible Factor-1 and iron regulatory protein 1 in iron uptake induced by acute hypoxia.

BACKGROUND INFORMATION: Divalent metal transporter 1 (DMT1) and transferrin receptor (TfR1) are vital proteins for cellular iron uptake. These proteins have hypoxia-responsive elements (HREs) in their 5'-regulatory region, and they are regulated by hypoxia-inducible factor 1α (HIF-1α) transcriptionally under hypoxic condition. Besides, iron regulatory protein 1 (IRP1) regulates DMT1 and TfR1 by binding to iron-responsive elements (IREs) present in their mRNAs to control cellular iron homeostasis. RESULTS: Here, we explored the effect of acute hypoxia on iron uptake. Ferrous iron uptake was elevated by DMT1(+IRE) and TfR1 under acute hypoxia. The luciferase activity analysis revealed that the functional HREs of DMT1 and TfR1 were increased. However, their IREs-dependent luciferase activities were reduced simultaneously. The mRNA stability of TfR1 and DMT1(+IRE) was suppressed under acute hypoxia. The mRNA levels of TfR1 and DMT1(+IRE) were restrain by silencing IRP1. In sharp contrast, HIF-1α overexpression enhanced the mRNA levels of TfR1 and DMT1(+IRE), which reversed the inhibition of IRP1 on both. HIF-1α konckdown suppressed the hypoxia-induced increase expression of TfR1 and DMT1(+IRE), whereas both proteins had little change when further decreased the IRP1 expression under hypoxia. Hypoxia upregulated the protein expression of Ferrtin-L in a time-dependent manner, yet there was no different when IRP1 silencing or overexperssion under hypoxia. The lactate dehydrogenase (LDH) release induced by hypoxia was increased by TfR1 siRNA silence. CONCLUSIONS: We propose that HIF-1/HRE system might play a principal part in hypoxia induced iron uptake.

A novel splicing site IRP1 somatic mutation in a patient with pheochromocytoma and JAK2V617F positive polycythemia vera: a case report.

BACKGROUND: The role of the hypoxia signaling pathway in the pathogenesis of pheochromocytoma/paraganglioma (PPGL)-polycythemia syndrome has been elucidated. Novel somatic mutations in hypoxia-inducible factor type 2A (HIF2A) and germline mutations in prolyl hydroxylase type 1 and type 2 (PHD1 and PHD2) have been identified to cause upregulation of the hypoxia signaling pathway and its target genes including erythropoietin (EPO) and its receptor (EPOR). However, in a minority of patients presenting with this syndrome, the genetics and molecular pathogenesis remain unexplained. The aim of the present study was to uncover novel genetic causes of PPGL-polycythemia syndrome. CASE PRESENTATION: A female presented with a history of JAK2V617F positive PV, diagnosed in 2007, and right adrenal pheochromocytoma diagnosed and resected in 2011. Her polycythemia symptoms and hematocrit levels continued to worsen from 2007 to 2011, with an increased frequency of phlebotomies. Postoperatively, until early 2013, her hematocrit levels remained normalized. Following this, the hematocrit levels ranged between 46.4 and 48.9% [35-45%]. Tumor tissue from the patient was further tested for mutations in genes related to upregulation of the hypoxia signaling pathway including iron regulatory protein 1 (IRP1), which is a known regulator of HIF-2α mRNA translation. Functional studies were performed to investigate the consequences of these mutations, especially their effect on the HIF signaling pathway and EPO. Indel mutations (c.267-1_267delGGinsTA) were discovered at the exon 3 splicing site of IRP1. Minigene construct and splicing site analysis showed that the mutation led to a new splicing site and a frameshift mutation of IRP1, which caused a truncated protein. Fluorescence in situ hybridization analysis demonstrated heterozygous IRP1 deletions in tumor cells. Immunohistochemistry results confirmed the truncated IRP1 and overexpressed HIF-2α, EPO and EPOR in tumor cells. CONCLUSIONS: This is the first report which provides direct molecular genetic evidence of association between a somatic IRP1 loss-of-function mutation and PHEO and secondary polycythemia. In patients diagnosed with PHEO/PGL and polycythemia with negative genetic testing for mutations in HIF2A, PHD1/2, and VHL, IRP1 should be considered as a candidate gene.

Investigating the role of the human CIA2A-CIAO1 complex in the maturation of aconitase.

BACKGROUND: The CIA2A protein, in complex with CIAO1, has been proposed to be exclusively implicated in the maturation of cytosolic aconitase. However, how the CIA2A-CIAO1 complex generates active aconitase is still unknown and the available structural information has not provided any crucial insights into the molecular function of CIA2A. METHODS: In this work we have characterized the Fe/S cluster binding properties of CIA2A and of the CIA2A-CIAO1 complex via NMR, UV - vis absorption and EPR spectroscopies and we have investigated how the Fe/S cluster is transferred to inactive aconitase/IRP1 protein. RESULTS: We found that an heterotrimeric species formed by two molecules of CIA2A and one of CIAO1 can bind one [4Fe-4S] cluster and that residue Cys90 of CIA2A is one of the cluster ligand. The holo trimeric complex is able to transfer the [4Fe-4S] cluster to apo-IRP1 thus generating the active form of aconitase. CONCLUSIONS AND GENERAL SIGNIFICANCE: These findings, which highlight a functional role for CIA2A-CIAO1 complex in aconitase maturation, raises a broad interest and can have a high impact on the community studying metal trafficking and iron­sulfur protein biogenesis. The present study can provide solid bases for further investigation of the molecular mechanisms involving also other CIA machinery proteins.

Evaluation of the iron regulatory protein-1 interactome.

The interactions of iron regulatory proteins (IRPs) with mRNAs containing an iron-responsive element (IRE) is a major means through which intracellular iron homeostasis is maintained and integrated with cellular function. Although IRE-IRP interactions have been proposed to modulate the expression of a diverse number of mRNAs, a transcriptome analysis of the interactions that form within the native mRNA structure and cellular environment has not previously been described. An RNA-CLIP study is described here that identified IRP-1 interactions occurring within a primary cell line expressing physiologically relevant amounts of mRNA and protein. The study suggests that only a small subset of the previously proposed IREs interact with IRP-1 in situ. Identifying authentic IRP interactions is not only important to a greater understanding of iron homeostasis and its integration with cell biology but also to the development of novel therapeutics that can compensate for iron imbalances.

Circadian Clock in a Mouse Colon Tumor Regulates Intracellular Iron Levels to Promote Tumor Progression.

Iron is an important biological catalyst and is critical for DNA synthesis during cell proliferation. Cellular iron uptake is enhanced in tumor cells to support increased DNA synthesis. Circadian variations in DNA synthesis and proliferation have been identified in tumor cells, but their relationship with intracellular iron levels is unclear. In this study, we identified a 24-h rhythm in iron regulatory protein 2 (IRP2) levels in colon-26 tumors implanted in mice. Our findings suggest that IRP2 regulates the 24-h rhythm of transferrin receptor 1 (Tfr1) mRNA expression post-transcriptionally, by binding to RNA stem-loop structures known as iron-response elements. We also found thatIrp2mRNA transcription is promoted by circadian clock genes, including brain and muscle Arnt-like 1 (BMAL1) and the circadian locomotor output cycles kaput (CLOCK) heterodimer. Moreover, growth in colon-26(Δ19) tumors expressing the clock-mutant protein (CLOCK(Δ19)) was low compared with that in wild-type colon-26 tumor. The time-dependent variation of cellular iron levels, and the proliferation rate in wild-type colon-26 tumor was decreased by CLOCK(Δ19)expression. Our findings suggest that circadian organization contributes to tumor cell proliferation by regulating iron metabolism in the tumor.

Cell death induced by mitochondrial complex I inhibition is mediated by Iron Regulatory Protein 1.

Mitochondrial dysfunction and oxidative damage, often accompanied by elevated intracellular iron levels, are pathophysiological features in a number of neurodegenerative processes. The question arises as to whether iron dyshomeostasis is a consequence of mitochondrial dysfunction. Here we have evaluated the role of Iron Regulatory Protein 1 (IRP1) in the death of SH-SY5Y dopaminergic neuroblastoma cells subjected to mitochondria complex I inhibition. We found that complex I inhibition was associated with increased levels of transferrin receptor 1 (TfR1) and iron uptake transporter divalent metal transporter 1 (DMT1), and decreased levels of iron efflux transporter Ferroportin 1 (FPN1), together with increased 55Fe uptake activity and an increased cytoplasmic labile iron pool. Complex I inhibition also resulted in increased oxidative modifications and increased cysteine oxidation that were inhibited by the iron chelators desferoxamine, M30 and Q1. Silencing of IRP1 abolished the rotenone-induced increase in 55Fe uptake activity and it protected cells from death induced by complex I inhibition. IRP1 knockdown cells presented higher ferritin levels, a lower iron labile pool, increased resistance to cysteine oxidation and decreased oxidative modifications. These results support the concept that IRP1 is an oxidative stress biosensor that mediates iron accumulation and cell death when deregulated by mitochondrial dysfunction. IRP1 activation, secondary to mitochondrial dysfunction, may underlie the events leading to iron dyshomeostasis and neuronal death observed in neurodegenerative disorders with an iron accumulation component.

Ferritin, cellular iron storage and regulation.

Ferritin is considered the major iron storage protein which maintains a large iron core in its cavity and has ferroxidase activity. There are many types of ferritin particularly in prokaryotes that include the canonical 24-mer FTN molecules, the heme-containing BFR, the smaller 12-mer DPS and the newly recognized EncFtn of encapsulin that forms a very large iron storage compartment. Recent studies show that ferritin function is more dynamic than previous depicted and new mechanisms of ferritin iron recycling are emerging. They participate to the regulation of cellular iron homeostasis as those of ferritin biosynthesis, cooperating also with the iron-dependent mechanism of cellular iron secretion. Some of these basic processes are in common between unicellular and animal cells, and this review aims at discussing the findings on the connections between iron storage, cellular iron regulation and ferritin iron recycling that have been explored in unicellular organisms and in animals. © 2017 IUBMB Life, 69(6):414-422, 2017.

Systemic iron homeostasis and erythropoiesis.

Iron is an essential nutrient that is potentially toxic due to its redox reactivity. Insufficient iron supply to erythroid cells, the major iron consumers in the body, leads to various forms of anemia. On the other hand, iron overload (hemochromatosis) is associated with tissue damage and diseases of liver, pancreas, and heart. Physiological iron balance is tightly controlled at the cellular and systemic level by iron regulatory proteins (IRP1, IRP2) and the iron regulatory hormone hepcidin, respectively. Underlying mechanisms often intersect to achieve optimal iron utilization, to control immune responses, and to prevent iron toxicity. This review focuses on systemic iron homeostasis in the context of erythropoiesis, a highly iron-demanding process. We discuss the function and regulation of hepcidin by various stimuli, and highlight hepcidin-dependent and -independent mechanisms that link iron utilization with maturation of erythroid progenitor cells. © 2017 IUBMB Life, 69(6):399-413, 2017.

Increased duodenal iron absorption through up-regulation of divalent metal transporter 1 from enhancement of iron regulatory protein 1 activity in patients with nonalcoholic steatohepatitis.

UNLABELLED: Increased hepatic iron accumulation is thought to be involved in the pathogenesis of nonalcoholic steatohepatitis (NASH). Hepatic iron accumulation, as well as oxidative DNA damage, is significantly increased in NASH livers. However, the precise mechanism of iron accumulation in the NASH liver remains unclear. In this study, 40 cases with a diagnosis of NASH (n = 25) or simple steatosis (SS; n = 15) by liver biopsy were enrolled. An oral iron absorption test (OIAT) was used, in which 100 mg of sodium ferrous citrate was administered to each individual. The OIAT showed that absorption of iron from the gastrointestinal (GI) tract was increased significantly in NASH patients, compared to SS and control subjects. Iron reduction therapy was effective in patients with NASH, who exhibited iron deposition in the liver and no alanine aminotransferase improvement after other therapies (n = 9). Serum hepcidin concentration and messenger RNA (mRNA) levels of divalent metal transporter 1 (DMT1) also were significantly elevated in patients with NASH. OIAT results were correlated with grade of liver iron accumulation and DMT1 mRNA levels. Then, we demonstrated that DMT1 mRNA levels increased significantly in Caco-2/TC7 cell monolayers cultured in transwells with serum from NASH patients. An electrophoresis mobility shift assay showed activation of iron regulatory protein (IRP) in those cells, and IRP1 small interfering RNA clearly inhibited the increase of DMT1 mRNA levels. CONCLUSION: In spite of elevation of serum hepcidin, iron absorption from the GI tract increased through up-regulation of DMT1 by IRP1 activation by humoral factor(s) in sera of patients with NASH.