Mechanism Of Iron Transport Research Articles

Regulation of plant iron homeostasis by abscisic acid: a review

Iron (Fe) is an important trace element involved in many important plant physiological and metabolic processes such as photosynthesis, respiration and nitrogen metabolism. Plants maintain iron homeostasis through absorption, transporting, storage and redistribution of iron. Iron metabolism is strictly regulated in plants. Iron regulatory transcription factors and iron transporters constitute the regulatory network of plant iron absorption and transport in plants. Ferritin and iron transporter jointly regulate the response to excess iron in plants. In recent years, important progress has been made in understanding how abscisic acid (ABA) regulates iron metabolism in plants. ABA may be used as a signal to regulate the absorption, transportation and reuse of Fe, or to relieve the symptoms of iron stress by regulating the oxidative stress responses in plants. In order to gain deeper insights into the crosstalk of ABA and iron metabolism in plants, this review summarized the mechanisms of iron absorption and transport and metabolic regulatory network in plants, as well as the mechanisms of ABA in regulating iron metabolism. The relationship between ABA and FER-like iron deficiency-induced transcription factor (FIT), iron-regulated transporter 1 (IRT1), and oxidative stress of iron deficiency were highlighted, and future research directions were prospected.

New insights into the mechanism of iron transport through the bacterial Ftr system present in pathogens.

Iron is an essential nutrient in bacteria. Its ferrous form, mostly present in low oxygen and acidic pH environments, can be imported using the specific Ftr-type transport system, which encompasses the conserved FtrABCD system found in pathogenic bacteria such as Bordetella, Brucella and Burkholderia. The nonpathogenicity and versatile metabolism of Rubrivivax gelatinosus make it an ideal model to study the FtrABCD system. Here, we report a new aspect of its regulation and the role of the periplasmic proteins FtrA and FtrB using in vivo and in vitro approaches. We investigated the metal binding mode and redox state of copper and iron to FtrA by crystallography and biophysical methods. An 'as isolated' FtrA protein from the bacterial periplasm contained a copper ion (Cu+ ) identified by electron paramagnetic resonance (EPR). Copper is coordinated by four conserved side chains (His and Met) in the primary metal site. Structural analysis of R. gelatinosus FtrA and FtrA homologues revealed that copper binding induces a rearrangement of the His95 imidazole ring, releasing thereafter space, as well as both Asp45 and Asp92 side chains, for iron binding in the secondary metal site. EPR highlighted that FtrA can oxidize the bound ferrous ion into the ferric form by reducing the bound Cu2+ into Cu+ , both metal sites being separated by 7 Å. Finally, we showed that FtrB binds iron and not copper. These results provide new insights into the mechanism of ferrous iron utilization by the conserved FtrABCD iron transporter for which we propose a new functional model.

Organic complexation of iron by strong ligands and siderophores in the eastern tropical North Pacific oxygen deficient zone

Continental margins are an important external source of dissolved iron to the marine environment. However, the mechanisms responsible for the offshore transport of dissolved iron is impacted by the resulting iron speciation. We characterized the iron speciation in the Eastern Tropical North Pacific (ETNP) oxygen deficient zone (ODZ), including dissolved iron, organic iron-binding ligands, and reduced iron. Organic iron-binding ligands were measured using both competitive ligand exchange adsorptive cathodic stripping voltammetry (CLE-ACSV) and liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS) in order to explore the impact of organic ligands on dissolved iron (dFeT) and iron(II) biogeochemistry in the region. Organic ligands were present in high concentrations (1.06–5.30 nmol L−1) and exceeded dissolved iron concentrations (0.36–4.52 nmol L−1) at all locations. Iron-binding strengths (logKFeL,Fe′cond) ranged 11.22 to 12.75 and were elevated in the ODZ layer relative to the oxygenated water column. LC-ESI-MS revealed the presence of siderophores, or bacterially-produced organic ligands with high Fe-affinity, in all samples analyzed, suggesting these compounds may be produced by microbes in the ODZ despite high ambient dFeT concentrations. This study is the first to characterize siderophores in an ODZ environment to date, and the three siderophores found (amphibactin B, synechobactin c9, synechobactin c10) could contribute to the observed elevated logKFeL,Fe′cond of ligands in the ODZ. Comparative analysis of organic ligand logKFeL,Fe′cond values in other low oxygen environments suggests that strong ligands, including siderophores, could be present in other low oxygen regions. In a simple model of the shelf-to-offshore iron transport mechanism, strong organic iron-binding ligands had a large impact on the longevity and transport of iron in the ODZ. These results suggest that organic ligand composition can have an impact on iron distributions in the ETNP ODZ and regulate the offshore transport of iron to the open ocean.

Quantification and comparison of gene expression associated with iron regulation and metabolism in a virulent and attenuated strain of Flavobacterium psychrophilum.

Iron is essential for growth and virulence in most pathogenic bacterial strains. In some cases, the hosts for these pathogenic bacteria develop specialized strategies to sequester iron and limit infectivity. This in turn may result in the invading pathogens utilizing high-affinity iron transport mechanisms, such as the use of iron-chelating siderophores, to extend beyond the host defences. Flavobacterium psychrophilum, the causative agent of bacterial coldwater disease (BCWD) in salmonids, relies on iron metabolism for infectivity, and the genome of the model CSF-259-93 strain has recently been made available. Further, this strain serves as a parent strain for a live-attenuated vaccine strain, B.17, which has been shown to provide rainbow trout with protection against BCWD. To elucidate specific gene expression responses to iron metabolism and compare strain differences, both F.psychrophilum strains were grown under iron-limiting conditions and 26 genes related to iron metabolism were mapped for 96hr in culture via qPCR analyses. Results indicate increased production of the ferrous iron transport protein B (FITB; p=.008), and ferric receptor CfrA (FR 1; p=.012) in the wild-type CSF-259-93 strain at 72hr and 96hr post-exposure to iron-limiting media. In the B.17 vaccine strain, siderophore synthase (SS) expression was found to be downregulated at 72hr, in comparison with 0h (p=.018). When strains were compared, FITB (p=.021), FR1 (p=.009) and SS (p=.016) were also elevated in B.17 at 0hr and TonB outer protein membrane receptor 1 (TBomr1; p=.005) had a lower expression at 96hr. Overall, this study demonstrated strain-related gene expression changes in only a fraction of the iron metabolism genes tested; however, results provide insight on potential virulence mechanisms and clarification on iron-related gene expression for F.psychrophilum.

10. Mechanism of iron transport and its regulation in plants

10. Mechanism of iron transport and its regulation in plants

Vibrio cholerae FeoB hydrolyzes ATP and GTP in vitro in the absence of stimulatory factors.

Feo is the most widely conserved system for ferrous iron transport in prokaryotes, and it is important for virulence in some pathogens. However, its mechanism of iron transport is not fully understood. In this study, we used full-length Vibrio cholerae FeoB (VcFeoB) as a model system to study whether its enzymatic activity is affected by regulatory factors commonly associated with FeoB proteins from other species or with G-proteins that have homology to FeoB. VcFeoB showed a higher rate of hydrolysis of both ATP and GTP than its N-terminal domain alone; likewise, ions such as K+ and Fe2+ did not modulate its nucleotide hydrolysis. We also showed that the three V. cholerae Feo proteins (FeoA, FeoB, and FeoC) work in a 1 : 1 : 1 molar ratio in vivo. Although both FeoA and FeoC are required for Feo-mediated iron transport, neither of these proteins affected the VcFeoB NTPase rate. These results are consistent with an active transport mechanism independent of stimulatory factors and highlight the importance of using full-length FeoB proteins as a reliable proxy to study Feo-mediated iron transport in vitro.

4096 Refined structure of human ferroportin using restraints from mass spectrometry

OBJECTIVES/GOALS: Mammals require iron for hemoglobin, respiration, immunity and as cofactor in enzymes. But free iron is toxic from the production of reactive oxygen species. Ferroportin is the sole exporter of cellular iron and it crucially determines cellular and systemic iron levels. Labile iron must be tightly regulated. This requires structural understanding. METHODS/STUDY POPULATION: We built structure of human ferroportin (FPN1) using the ab ignition prediction approaches of Rosetta/Robetta and by comparative modeling with distance restraints in MODELLER. Templates selected were from solute carrier protein families of distantly related orthologs and homologs including a proton coupled peptide transporter (PDB ID: 4IKV) and the bacterial iron transporter in outward-open and inward-open states, (PDB ID: 5AYM, 5AYO). Each model was validated by experimental mass spectrometry data. The energy minimized structural model was inserted into a lipid bilayer, placed in a rectangular simulation box, covered with TIP3P water solvent balanced with counterions and conditioned. Finally, we carried out 350 nanoseconds molecular dynamics simulations. RESULTS/ANTICIPATED RESULTS: Our first model of FPN1 (571aa), using Rosetta/Robetta ab initio approach, resembles the structure of the proton-dependent transporter, POT and consists of 12 transmembrane helices. The membrane spanning helices veer away from the orientation in the structure of 4IKV. The alternate model using MODELLER and the method of satisfaction of constraints, returned one template, the structure of Bdellovibrio bacteriovorus iron (Fe2+) transporter homolog (5AYN, 440aa) with sequence identity of 19%. Aligning FPN1 on the template sequence incorporating structural information revealed better conservation (29%). This model also comprises 12 transmembrane helices in two bundles separated by a large intracellular loop. The iron binding site predicted in both models match the structures of distant bacterial homologs. DISCUSSION/SIGNIFICANCE OF IMPACT: We are using these experimentally verified structures and functional data to answer questions about the mechanism of ferroportin iron transport, structural dynamics and the significance of mutations in ferroportin seen in different populations, especially the Q248H mutation found in Africans and black Americans with moderate to high prevalence.

Ferric Chelate Reductase Knockdown in Drosophila S2 Cells

Although the mechanisms of cellular iron uptake in mammals are largely understood, there is still little known about the mechanism of iron transport in insects. Not only is iron essential, it can also be toxic and therefore needs to be well regulated to maintain balance between necessity and toxicity. In mammals, a duodenal cytochrome b acts as a ferric reductase to allow for uptake of iron into intestinal cells. We identified a cytochrome b561 family member in Drosophila, CG8399, that is expressed in Drosophila S2 cells. CG8399 is similar to ferric chelate reductase‐1 in humans as both contain reeler, domon and cytochrome b561 domains. We are testing the hypothesis that there is reduction of iron by CG8399, which allows the transport of iron into the cells. To test this hypothesis, we used RNA interference (RNAi) to knockdown the CG8399 mRNA. We quantified the RNAi knockdown caused by RNAi using real time PCR and compared iron concentrations between untreated and treated cells using a ferrozine‐based iron content assay. A lower concentration of iron in CG8399 RNAi‐treated cells would support our hypothesis.Support or Funding InformationNSF IOS‐1656407

Secreted insect transferrin‐1 with strong and reversible iron‐binding has potentially tissue specific roles in immunity and iron transport

Iron is biologically essential and potentially toxic. To maintain healthy physiological systems, iron must be properly sequestered, transported and stored in animals. The transferrin family of proteins plays a major role in maintaining this homeostasis in vertebrate animals. However, little is understood about iron homeostasis and transferrin’s role in invertebrates. In mammals there are two well understood secreted iron‐binding transferrins: (1) serum transferrin (STF) binds two ferric‐iron ions in the blood and transports them into cells by receptor mediate endocytosis; (2) lactoferrin (LTF) functions as an immune protein by sequestering two ferric‐iron ions in secreted fluids to inhibit bacterial growth. Insect Transferrin‐1 (Tsf1) is found in insect hemolymph and secreted fluids and is homologous to STF and LTF; however, its function in the iron transport and immunity of invertebrates is not clear. As a first step towards elucidating its function, I have purified recombinant Drosophila melanogaster Tsf1 (DmTsf1) and hemolymph derived Manduca sexta Tsf1 (MsTsf1). Comparing the spectroscopic and kinetic data of DmTsf1 and MsTsf1 to STF and LTF shows that these Tsf1s coordinate iron differently and have a relatively lower but still potent iron binding affinity at pH 7.4 (DmTsf1 log K′= 17, MsTsf1 log K′= 17 and STF log K′ = 20). The Tsf1s release iron over the pH range of approximately 6 to 4, which is nearly identical to STF and indicates the possibility of iron release in the endosome. Current work is aimed toward determining how iron from Tsf1 is transported into cells. One hypothesis is that Tsf1 is endocytosed and iron is released in the endosome—similar to the iron transport mechanism of STF in mammals. I tested this hypothesis using immunohistochemistry and confocal imaging of Drosophila tissues. Immunostaining shows that DmTsf1 colocalizes with an early endosome marker (Rab5) in the developing oocyte, supporting my hypothesis of iron delivery. DmTsf1 was also found in the tracheal tissue, but instead of clearly colocalizing with Rab5, it appears that Tsf1 is secreted into the tracheal lumen and not taken back up. Overall, these results indicate that Tsf1s may have tissue specific roles: iron delivery to the oocyte and iron sequestration in the tracheal lumen. Future work will be completed to test if Tsf1 is endocytosed in other tissues and if it plays an important role in supplying iron to these tissues.Support or Funding InformationNSF: 1656388

Molecular model of the ferroportin intracellular gate and implications for the human iron transport cycle and hemochromatosis type 4A.

Ferroportin 1 (FPN1) is a major facilitator superfamily transporter that is essential for proper maintenance of human iron homeostasis at the systemic and cellular level. FPN1 dysfunction leads to the progressive accumulation of iron in reticuloendothelial cells, causing hemochromatosis type 4A (or ferroportin disease), an autosomal dominant disorder that displays large phenotypic heterogeneity. Although crystal structures have unveiled the outward- and inward-facing conformations of the bacterial homolog Bdellovibrio bacteriovorus Fpn (or Bd2019) and calcium has recently been identified as an essential cofactor, our molecular understanding of the iron transport mechanism remains incomplete. Here, we used a combination of molecular modeling, molecular dynamics simulations, and Ala site-directed mutagenesis, followed by complementary in vitro functional analyses, to explore the structural architecture of the human FPN1 intracellular gate. We reveal an interdomain network that involves 5 key amino acids and is likely very important for stability of the iron exporter facing the extracellular milieu. We also identify inter- and intradomain interactions that rely on the 2 Asp84 and Asn174 critical residues and do not exist in the bacterial homolog. These interactions are thought to play an important role in the modulation of conformational changes during the transport cycle. We interpret these results in the context of hemochromatosis type 4A, reinforcing the idea that different categories of loss-of-function mutations exist. Our findings provide an unprecedented view of the human FPN1 outward-facing structure and the particular function of the so-called "gating residues" in the mechanism of iron export.-Guellec, J., Elbahnsi, A., Le Tertre, M., Uguen, K., Gourlaouen, I., Férec, C., Ka, C., Callebaut, I., Le Gac, G. Molecular model of the ferroportin intracellular gate and implications for the human iron transport cycle and hemochromatosis type 4A.

The multifaceted role of iron in renal health and disease.

Iron is an essential element that is indispensable for life. The delicate physiological body iron balance is maintained by both systemic and cellular regulatory mechanisms. The iron-regulatory hormone hepcidin assures maintenance of adequate systemic iron levels and is regulated by circulating and stored iron levels, inflammation and erythropoiesis. The kidney has an important role in preventing iron loss from the body by means of reabsorption. Cellular iron levels are dependent on iron import, storage, utilization and export, which are mainly regulated by the iron response element-iron regulatory protein (IRE-IRP) system. In the kidney, iron transport mechanisms independent of the IRE-IRP system have been identified, suggesting additional mechanisms for iron handling in this organ. Yet, knowledge gaps on renal iron handling remain in terms of redundancy in transport mechanisms, the roles of the different tubular segments and related regulatory processes. Disturbances in cellular and systemic iron balance are recognized as causes and consequences of kidney injury. Consequently, iron metabolism has become a focus for novel therapeutic interventions for acute kidney injury and chronic kidney disease, which has fuelled interest in the molecular mechanisms of renal iron handling and renal injury, as well as the complex dynamics between systemic and local cellular iron regulation.

Ferric Reductase Knockdown in Drosophila S2 Cells

Iron is an essential micronutrient that is potentially harmful due to its redox properties, which is why its transport between cells must be a highly regulated process. As compared to mammals, very little is known about iron uptake in insects, because receptors for ferritin or transferrin have not been identified in insects. The goal of the proposed research is to understand the physiological mechanisms of iron transport in insects. We are testing the hypothesis that the reduction of iron ions in the hemolymph leads to iron transport into cells. Hemolymph contains iron in the ferric form, but iron transporters in animal cells are specific to ferrous iron; therefore iron uptake via an influx transporter is likely to require a reduction step. We silenced a putative ferric reductase CG8399 by RNA interference (RNAi) to determine if it contributes to ferric reductase activity and impacts the iron content of the cells. We analyzed the efficiency of knockdown of CG8399 by real time, quantitative PCR (qPCR).Support or Funding InformationNSF IOS‐1656407This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Polymorphism of the Iron Homeostasis Genes and Iron Sensitivity in Saccharomyces cerevisiae Flor and Wine Strains

Iron is an essential micronutrient for all living organisms. The mechanisms of iron transport and homeostasis have been studied in detail in Saccharomyces cerevisiae yeasts, and iron metabolism in the strains isolated from different ecological niches was found to vary. In the present work, comparative analysis of iron homeostasis in the wine and flor yeast strains was carried out. Flor yeasts are characterized by their ability to grow as a biofilms on the surface of fortified wine materials, where they carry out oxidative metabolism. We have found that flor strains are more sensitive to elevated iron concentrations as compared to wine strains and accumulate iron more efficiently when grown in iron-limited media. These features correlate with the Q648X mutation in the Aft1p transcription factor, resulting in emergence of its truncated form and with the chromosomal rearrangement resulting in the deletion of the FRE-FIT cluster containing the Fre3p and Fre5p iron reductase genes and of the Fit2p and Fit3p membrane proteins responsible for accumulation of siderophore-bound iron in the cell wall. Our results indicate a significant adaptive role of these variations in the iron homeostasis genes for the physiology of flor yeasts, since selection of these organisms was accompanied by adaptation to growth on materials with low iron content, such as fortified fermented wort obtained from grapes grown in traditional areas of sherry wine-making, which is important for the preservation of specific properties of sherry wines.

Emerging Mechanisms of Cellular Iron Transport and Trafficking

Iron is an essential nutrient required by every cell in the human body, yet it can also be a potent cellular toxin. Iron is essential because enzymes that require iron co-factors (namely, heme, iron-sulfur clusters, mono- and dinuclear iron centers) are involved in virtually every major metabolic process in the cell. Iron deficiency continues to be the most common nutritional deficiency in the world, while iron overload is a feature of an increasing number of human diseases, including genetic disorders such as hereditary hemochromatosis, thalassemias, and Friedreich ataxia, as well as chronic inflammatory diseases of the liver, such as hepatitis C and steatohepatitis. The biochemical mechanisms by which iron causes toxicity are not completely known. Previously, we identified Poly rC-binding protein 1 (Pcbp1) as a protein that directly binds and delivers iron to ferritin, the major iron storage protein in mammalian cells. Pcbp1 and its paralog Pcbp2 are multifunctional adaptors that bind cellular RNA, DNA, proteins, and iron, altering the fate of their binding partners. Ferritin is a large polypeptide, containing 24 subunits of H- and L-chains assembled into a hollow sphere. Reduced (ferrous) iron enters the sphere through pores formed between the subunits and is oxidized on the interior surface to form nanocrystals of ferric oxyhydroxides. Pcbp1 is the first example of an iron chaperone- a protein that specifically binds iron ions and delivers them to target proteins, such as ferritin, through direct protein-protein interactions. In cultured cells, Pcbp1 is also required for efficient iron delivery to non-heme enzymes in addition to ferritin. These enzymes include the mononuclear iron-containing prolyl hydroxylases that regulate HIF and the dinuclear iron-containing deoxyhypusine hydroxylase, which is required for the modification of lysine to hypusine. In the adult human, 70% of total body iron is present in circulating erythrocytes, which are produced by the bone marrow at a rate of 2 million reticulocytes per second. We examined the roles of Pcbp1 and Ncoa4 in this extraordinary flux of iron through the erythron. Ncoa4 is the cargo receptor that recruits ferritin into the autophagosome for degradation in the lysosome. Using an in vitro model of erythroid differentiation, we showed that depletion of Pcbp1 or Ncoa4 impeded trafficking of iron through ferritin, which impaired the synthesis of heme and hemoglobin. Mice with tamoxifen-induced Pcbp1 deficiency exhibited a microcytic anemia typical of iron deficiency, with activation of compensatory erythropoiesis. The role of ferritin in erythropoiesis has been controversial, but our studies indicate that iron flux through ferritin is an obligatory process in the early stages of erythroid terminal differentiation. Our in vitro studies of erythrocytes revealed that the interactions of Pcbp1 and Ncoa4 with ferritin changed during differentiation, with maximal binding of Pcbp1 at early stages and Ncoa4 at late stages. These binding activities are regulated by cellular iron in mechanistically distinct ways. Ongoing studies of Pcbp1 deficiency in other murine tissues suggest the importance of iron chaperones in maintaining the bioavailable pool of iron in mammalian cells. DisclosuresNo relevant conflicts of interest to declare.

Influence of Physical Activity on the Regulation of Iron Metabolism

Intense physical activity has a significant effect on iron metabolism in the human organism. This paper is a review of world literature data focused on the mechanisms of iron metabolism regulation in the human organism, as well as the influence of intense exercise on these regulatory mechanisms. We have discussed the mechanisms of iron absorption and transport and the regulation of these processes under normal conditions. It has been found that intense exercise is accompanied by increased production of interleukin-6, a positive regulator of hepcidin production. Hepcidin, in turn, has an inhibitory effect on the expression of divalent metal transporter 1 and ferroportin, finally leading to both decreased iron absorption and iron sequestration in cells. All of the mentioned stages of iron deficiency development may be promising targets for correction of iron balance and working ability in patients exposed to intense physical activity.

Homology modelling of human divalent metal transporter (DMT): Molecular docking and dynamic simulations for duodenal iron transport

Iron transport through the duodenum is regulated by carrier proteins, one of which is the ubiquitously distributed divalent metal transporter (DMT1) which is responsible for the uptake of iron across the apical surface of the duodenal enterocyte. The crystallographic structure of Staphylococcus capitis divalent metal ion transporter (ScaDMT1) was obtained and it was used as a template for the construction of a homology model of human divalent metal transporter (hDMT1). The binding site for hDMT1 was determined by using SiteMap as well as molecular docking studies on ScaDMT1. The differences in binding modes between ScaDMT1 and hDMT1 were noted for a set of 7 iron containing compounds, including ferrous sulphate. Diffusion of ferrous ion was observed during the course of molecular dynamic simulation which corresponded to the postulated mechanism of iron transport. Further, the dock scores correlated well with relative bioavailabilities of the iron compounds. The study confirmed the efficacy of the in silico model which could be used for future studies on the absorption of micronutrients.

Bioinorganic Chemistry of Iron

The X-ray crystallographic analysis of the single-crystal mugineic acid-Cu(II) complex showed that mugineic acid acts as a hexadentate ligand. Mugineic acid, a typical phytosiderophore, shows a marked stimulating effect on 59Fe-uptake and chlorophyll synthesis in rice plants. A salient feature is the higher reduction potential of the mugineic acid-Fe(III) complex than those of bacterial siderophores. X-ray diffraction study of the structurally analogous Co(III) complex of the mugineic acid-Fe(III) complex demonstrates that the azetidine nitrogen and secondary amine nitrogen, and both terminal carboxylate oxygens, coordinate as basal planar donors, and the hydroxyl oxygen and intermediate carboxylate oxygen bind as axial donors in a nearly octahedral configuration. The iron-transport mechanism in gramineous plants appears to involve the excretion of mugineic acid from the roots, which aids Fe(III)-solubilization and reduction of Fe(III) to Fe(II). Manganese peroxidase (MnP) is a component of the lignin degradation system of the basidiomycetous fungus, Phanerochaete chrysosporium. To elucidate the heme environment of this novel Mn(II)-dependent extracellular enzyme, we studied its ESR and resonance Raman spectroscopic properties. Consequently, it is most likely that the heme environment of MnP resembles that of cytochrome c peroxidase. In addition, degradation methods using basidiomycetous fungi or Fe3+-H2O2 mixed reagent were developed for dioxins and polychlorinated biphenyls. The complete amino acid sequences of respective [2Fe-2S] ferredoxins were determined and compared with those of other higher plants. Finally, the toxic effects of iron on human health and the development of novel antibacterial drugs capable of inhibiting the iron transport system of Vibrio vulnificus are described.

Regulatory mechanisms for iron transport across the blood-brain barrier

Many critical metabolic functions in the brain require adequate and timely delivery of iron. However, most studies when considering brain iron uptake have ignored the iron requirements of the endothelial cells that form the blood-brain barrier (BBB). Moreover, current models of BBB iron transport do not address regional regulation of brain iron uptake or how neurons, when adapting to metabolic demands, can acquire more iron. In this study, we demonstrate that both iron-poor transferrin (apo-Tf) and the iron chelator, deferoxamine, stimulate release of iron from iron-loaded endothelial cells in an in vitro BBB model. The role of the endosomal divalent metal transporter 1 (DMT1) in BBB iron acquisition and transport has been questioned. Here, we show that inhibition of DMT1 alters the transport of iron and Tf across the endothelial cells. These data support an endosome-mediated model of Tf-bound iron uptake into the brain and identifies mechanisms for local regional regulation of brain iron uptake. Moreover, our data provide an explanation for the disparity in the ratio of Tf to iron transport into the brain that has confounded the field.

Hypoxia and iron requirements are the main drivers in transcriptional adaptation of Kluyveromyces lactis during wine aerobic fermentation

The respiratory metabolism of yeast species alternative to Saccharomyces cerevisiae has been explored in recent years as a tool to reduce ethanol content in grape wine. The efficacy of this strategy has been previously proven for mixed cultures of non-Saccharomyces and S. cerevisiae strains. In this work, we perform a transcriptomic analysis of the Crabtree-negative yeast Kluyveromyces lactis under tightly controlled growth conditions in order to better understand physiology of non-Saccharomyces yeasts during the fermentation of grape must under aerated conditions. Transcriptional changes in K. lactis are mainly driven by oxygen limitation, iron requirement, and oxidative stress. Oxidative stress appears as a consequence of the hypoxic conditions achieved by K. lactis once oxygen supply is no longer sufficient to sustain fully respiratory metabolism. This species copes with low oxygen and iron availability by repressing iron consuming pathways and activating iron transport mechanisms. Most of the physiological and transcriptomic features of K. lactis in aerobic wine fermentation are not shared with the Crabtree-positive yeast S. cerevisiae.

Identifying the Genes Responsible for Iron-Limited Condition in Riemerella anatipestifer CH-1 through RNA-Seq-Based Analysis.

One of the important elements for most bacterial growth is iron, the bioavailability of which is limited in hosts. Riemerella anatipestifer (R. anatipestifer, RA), an important duck pathogen, requires iron to live. However, the genes involved in iron metabolism and the mechanisms of iron transport are largely unknown. Here, we investigated the transcriptomic effects of iron limitation condition on R. anatipestifer CH-1 using the RNA-Seq and RNA-Seq-based analysis. Data analysis revealed genes encoding functions related to iron homeostasis, including a number of putative TonB-dependent receptor systems, a HmuY-like protein-dependent hemin (an iron-containing porphyrin) uptake system, a Feo system, a gene cluster related to starch utilization, and genes encoding hypothetical proteins that were significantly upregulated in response to iron limitation. Compared to the number of upregulated genes, more genes were significantly downregulated in response to iron limitation. The downregulated genes mainly encoded a number of outer membrane receptors, DNA-binding proteins, phage-related proteins, and many hypothetical proteins. This information suggested that RNA-Seq-based analysis in iron-limited medium is an effective and fast method for identifying genes involved in iron uptake in R. anatipestifer CH-1.