Divergent roles of IREG/Ferroportin transporters from the nickel hyperaccumulator Leucocroton havanensis.

In response to our ever-increasing demand for metals, phytotechnologies are being developed to limit the environmental impact of conventional metal mining. However, the development of these technologies, which rely on plant species able to tolerate and accumulate metals, is partly limited by our lack of knowledge of the underlying molecular mechanisms. In this work, we aimed to better understand the role of metal transporters of the IRON REGULATED 1/FERROPORTIN (IREG/FPN) family from the nickel hyperaccumulator Leucocroton havanensis from the Euphorbiaceae family. Using transcriptomic data, we identified two homologous genes, LhavIREG1 and LhavIREG2, encoding divalent metal transporters of the IREG/FPN family. Both genes are expressed at similar levels in shoots, but LhavIREG1 shows higher expression in roots. The heterologous expression of these transporters in A. thaliana revealed that LhavIREG1 is localized to the plasma membrane, whereas LhavIREG2 is located on the vacuole. In addition, the expression of each gene induced a significant increase in nickel tolerance. Taken together, our data suggest that LhavIREG2 is involved in nickel sequestration in vacuoles of leaf cells, whereas LhavIREG1 is mainly involved in nickel translocation from roots to shoots, but could also be involved in metal sequestration in cell walls. Our results suggest that paralogous IREG/FPN transporters may play complementary roles in nickel hyperaccumulation in plants.

Role of zinc in health and disease.

This review provides a concise overview of the cellular and clinical aspects of the role of zinc, an essential micronutrient, in human physiology and discusses zinc-related pathological states. Zinc cannot be stored in significant amounts, so regular dietary intake is essential. ZIP4 and/or ZnT5B transport dietary zinc ions from the duodenum into the enterocyte, ZnT1 transports zinc ions from the enterocyte into the circulation, and ZnT5B (bidirectional zinc transporter) facilitates endogenous zinc secretion into the intestinal lumen. Putative promoters of zinc absorption that increase its bioavailability include amino acids released from protein digestion and citrate, whereas dietary phytates, casein and calcium can reduce zinc bioavailability. In circulation, 70% of zinc is bound to albumin, and the majority in the body is found in skeletal muscle and bone. Zinc excretion is via faeces (predominantly), urine, sweat, menstrual flow and semen. Excessive zinc intake can inhibit the absorption of copper and iron, leading to copper deficiency and anaemia, respectively. Zinc toxicity can adversely affect the lipid profile and immune system, and its treatment depends on the mode of zinc acquisition. Acquired zinc deficiency usually presents later in life alongside risk factors like malabsorption syndromes, but medications like diuretics and angiotensin-receptor blockers can also cause zinc deficiency. Inherited zinc deficiency condition acrodermatitis enteropathica, which occurs due to mutation in the SLC39A4 gene (encoding ZIP4), presents from birth. Treatment involves zinc supplementation via zinc gluconate, zinc sulphate or zinc chloride. Notably, oral zinc supplementation may decrease the absorption of drugs like ciprofloxacin, doxycycline and risedronate.

Plasma non-transferrin-bound iron uptake by the small intestine leads to intestinal injury and intestinal flora dysbiosis in an iron overload mouse model and Caco-2 cells.

Iron overload often occurs during blood transfusion and iron supplementation, resulting in the presence of non-transferrin-bound iron (NTBI) in host plasma and damage to multiple organs, but effects on the intestine have rarely been reported. In this study, an iron overload mouse model with plasma NTBI was established by intraperitoneal injection of iron dextran. We found that plasma NTBI damaged intestinal morphology, caused intestinal oxidative stress injury and reactive oxygen species (ROS) accumulation, and induced intestinal epithelial cell apoptosis. In addition, plasma NTBI increased the relative abundance of Ileibacterium and Desulfovibrio in the cecum, while the relative abundance of Faecalibaculum and Romboutsia was reduced. Ileibacterium may be a potential microbial biomarker of plasma NTBI. Based on the function prediction analysis, plasma NTBI led to the weakening of intestinal microbiota function, significantly reducing the function of the extracellular structure. Further investigation into the mechanism of injury showed that iron absorption in the small intestine significantly increased in the iron group. Caco-2 cell monolayers were used as a model of the intestinal epithelium to study the mechanism of iron transport. By adding ferric ammonium citrate (FAC, plasma NTBI in physiological form) to the basolateral side, the apparent permeability coefficient (Papp) values from the basolateral to the apical side were greater than 3×10-6 cm s-1. Intracellular ferritin level and apical iron concentration significantly increased, and SLC39A8 (ZIP8) and SLC39A14 (ZIP14) were highly expressed in the FAC group. Short hairpin RNA (shRNA) was used to knock down ZIP8 and ZIP14 in Caco-2 cells. Transfection with ZIP14-specific shRNA decreased intracellular ferritin level and inhibited iron uptake. These results revealed that plasma NTBI may cause intestinal injury and intestinal flora dysbiosis due to the uptake of plasma NTBI from the basolateral side into the small intestine, which is probably mediated by ZIP14.

Comprehensive analysis of OpHD-ZIP transcription factors related to the regulation of camptothecin biosynthesis in Ophiorrhiza pumila.

Ophiorrhiza pumila, as a folk herb belonging to the Rubiaceae family, has become a potential source of camptothecin (CPT), which is a monoterpenoid indole alkaloid with good antitumor property. However, the camptothecin content in this herb is low, and is far from meeting the increasing clinical demand. Understanding the transcriptional regulation of camptothecin biosynthesis provides an effective strategy for improvement of camptothecin yield. Previous studies have demonstrated several transcription factors that are related to camptothecin biosynthesis, while the functions of HD-ZIP members in O. pumila have not been investigated yet. In this study, 32 OpHD-ZIP transcription factor members were genome-wide identified. Phylogenetic tree showed that these OpHD-ZIP proteins are divided into four subfamilies. Based on the transcriptome data, nine OpHD-ZIP genes were shown to be predominantly expressed in O. pumila roots, which were in line with the camptothecin biosynthetic genes. Co-expression analysis showed that OpHD-ZIP7 and OpHD-ZIP20 were potentially related to the modulation of camptothecin biosynthesis. Dual-luciferase reporter assays (Dual-LUC) showed that both OpHD-ZIP7 and OpHD-ZIP20 could activate the expression of camptothecin biosynthetic genes OpIO and OpTDC. In conclusion, this study offered the promising data for exploring the roles of OpHD-ZIP transcription factors in regulating camptothecin biosynthesis.

Insights into the regulation of cellular Mn2+ homeostasis via TMEM165.

Golgi cation homeostasis is known to be crucial for many cellular processes including vesicular fusion events, protein secretion, as well as for the activity of Golgi glycosyltransferases and glycosidases. TMEM165 was identified in 2012 as the first cation transporter related to human glycosylation diseases, namely the Congenital Disorders of Glycosylation (CDG). Interestingly, divalent manganese (Mn) supplementation has been shown to suppress the observed glycosylation defects in TMEM165-deficient cell lines, thus suggesting that TMEM165 is involved in cellular Mn homeostasis. This paper demonstrates that the origin of the Golgi glycosylation defects arises from impaired Golgi Mn homeostasis in TMEM165-depleted cells. We show that Mn supplementation fully rescues the Mn content in the secretory pathway/organelles of TMEM165-depleted cells and hence the glycosylation process. Strong cytosolic and organellar Mn accumulations can also be observed in TMEM165- and SPCA1-depleted cells upon incubation with increasing Mn concentrations, thus demonstrating the crucial involvement of these two proteins in cellular Mn homeostasis. Interestingly, our results show that the cellular Mn homeostasis maintenance in control cells is correlated with the presence of TMEM165 and that the Mn-detoxifying capacities of cells, through the activity of SPCA1, rely on the Mn-induced degradation mechanism of TMEM165. Finally, this paper highlights that TMEM165 is essential in secretory pathway/organelles Mn homeostasis maintenance to ensure both Golgi glycosylation enzyme activities and cytosolic Mn detoxification.

Enterocyte-specific deletion of metal transporter Zip14 (Slc39a14) alters intestinal homeostasis through epigenetic mechanisms.

Zinc has anti-inflammatory properties using mechanisms that are unclear. Zip14 (Slc39a14) is a zinc transporter induced by proinflammatory stimuli and is highly expressed at the basolateral membrane of intestinal epithelial cells (IECs). Enterocyte-specific Zip14 ablation (Zip14ΔIEC) in mice was developed to study the functions of this transporter in enterocytes. This gene deletion led to increased intestinal permeability, increased IL-6 and IFNγ expression, mild endotoxemia, and intestinal dysbiosis. RNA sequencing was used for transcriptome profiling. These analyses revealed differential expression of specific intestinal proinflammatory and tight junction (TJ) genes. Binding of transcription factors, including NF-κß, STAT3, and CDX2, to appropriate promoter sites of these genes supports the differential expression shown with chromatin immunoprecipitation assays. Total histone deacetylase (HDAC), and specifically HDAC3, activities were markedly reduced with Zip14 ablation. Intestinal organoids derived from ΔIEC mice display TJ and cytokine gene dysregulation compared with control mice. Differential expression of specific genes was reversed with zinc supplementation of the organoids. We conclude that zinc-dependent HDAC enzymes acquire zinc ions via Zip14-mediated transport and that intestinal integrity is controlled in part through epigenetic modifications.NEW & NOTEWORTHY We show that enterocyte-specific ablation of zinc transporter Zip14 (Slc39a14) results in selective dysbiosis and differential expression of tight junction proteins, claudin 1 and 2, and specific cytokines associated with intestinal inflammation. HDAC activity and zinc uptake are reduced with Zip14 ablation. Using intestinal organoids, the expression defects of claudin 1 and 2 are resolved through zinc supplementation. These novel results suggest that zinc, an essential micronutrient, influences gene expression through epigenetic mechanisms.

Modulation of Zinc Transporter Expressions by Additional Zinc in C2C12 Cells Cultured in a High Glucose Environment and in the Presence of Insulin or Interleukin-6.

Zn status has been related to various chronic diseases presenting oxidative stress and inflammation, such as type 2 diabetes. Zn supplementation has been suggested to be a potential coadjuvant in the management of this condition. Zn transporters constitute a key component in the maintenance of Zn homeostasis. Our aim was to evaluate the modulatory effect of additional Zn (10 or 100 µM; as a ZnSO4*7H20) on the mRNA relative expression of selected Zn transporters (ZnT1, ZnT5, ZnT7, ZIP6, ZIP7, ZIP10, ZIP14), in myoblast (C2C12) cells cultured in normal (10 mM) and high glucose (30 mM), and in the absence or presence of insulin (1 nM), and interleukin-6 (IL-6; 5 nM) for 24 h. The main findings of our study were that in high glucose conditions in absence of insulin or IL-6, additional Zn increased ZnT1 and ZIP6, and decreased ZnT5 and ZIP7 expressions. However, this situation is modified by insulin, where incremental Zn induced increased expressions of ZnT1, ZnT5, and all the ZIP transporters studied. In high glucose conditions and in the presence of IL-6, additional Zn caused increased expressions of ZnT7, ZIP7, and ZIP14, compared with results in the absence of IL-6. This study provides preliminary evidence for the differential expression of selected Zn transporters in C2C12 cells subjected to high glucose and incremental Zn, suggesting that important changes in intracellular Zn distribution take place in response to inflammatory and high-insulin environments. Further study is necessary to understand the implications of these findings.

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A girl, aged 11 years, was admitted due to recurrent rash on the whole body and mucosa for 10 years, and typical rash was erythema at the perioral region, hand-foot joints, vulva, and perianal region, with blisters, erosions, and ulcers on the erythema. The girl was improved after zinc supplementation. Her younger brother had similar rash and medical history. The histopathological examination showed epidermal parakeratosis with mild hyperkeratosis, severe spongiform edema of the stratum corneum, significant proliferation of acanthocytes, and vacuolation of keratinocytes. The genetic testing revealed that both the girl and her younger brother had a homozygous mutation of c.1456(exon9)delG in the SLC39A4 gene, and thus the girl was diagnosed with acrodermatitis enteropathica. It is concluded that for children with recurrent rash on the limbs and at the perioral region, genetic testing should be performed as early as possible to make a confirmed diagnosis, and a sufficient dose of zinc supplementation should be given, while the levels of trace elements such as blood zinc should be regularly monitored.

Deficiency in the zinc transporter ZIP8 impairs epithelia renewal and enhances lung fibrosis.

Although aging and lung injury are linked to the development of idiopathic pulmonary fibrosis (IPF), the underlying pathognomonic processes predisposing to fibrotic lesions remain largely unknown. A deficiency in the ability of type 2 alveolar epithelial cell (AEC2) progenitors to regenerate and repair the epithelia has been proposed as a critical factor. In this issue of the JCI, Liang et al. identify a deficiency in the zinc transporter SLC39A8 (ZIP8) in AEC2s and in the subsequent activation of the sirtuin SIRT1 that predisposes to decreased AEC2 renewal capacity and enhanced lung fibrosis in both IPF and aging lungs. Interestingly, the authors demonstrate the efficacy of modulating dietary zinc levels, suggesting the need for clinical trials to evaluate the therapeutic potential of dietary supplementation and the development of pharmacological modulation of the Zn/ZIP8/SIRT1 axis for treatment.

HKT1;1 and HKT1;2 Na+ Transporters from Solanum galapagense Play Different Roles in the Plant Na+ Distribution under Salinity.

Salt tolerance is a target trait in plant science and tomato breeding programs. Wild tomato accessions have been often explored for this purpose. Since shoot Na+/K+ is a key component of salt tolerance, RNAi-mediated knockdown isogenic lines obtained for Solanum galapagense alleles encoding both class I Na+ transporters HKT1;1 and HKT1;2 were used to investigate the silencing effects on the Na and K contents of the xylem sap, and source and sink organs of the scion, and their contribution to salt tolerance in all 16 rootstock/scion combinations of non-silenced and silenced lines, under two salinity treatments. The results show that SgHKT1;1 is operating differently from SgHKT1;2 regarding Na circulation in the tomato vascular system under salinity. A model was built to show that using silenced SgHKT1;1 line as rootstock would improve salt tolerance and fruit quality of varieties carrying the wild type SgHKT1;2 allele. Moreover, this increasing effect on both yield and fruit soluble solids content of silencing SgHKT1;1 could explain that a low expressing HKT1;1 variant was fixed in S. lycopersicum during domestication, and the paradox of increasing agronomic salt tolerance through silencing the HKT1;1 allele from S. galapagense, a salt adapted species.

Iron from nanostructured ferric phosphate: absorption and biodistribution in mice and bioavailability in iron deficient anemic women.

Food fortification with iron nanoparticles (NPs) could help prevent iron deficiency anemia, but the absorption pathway and biodistribution of iron-NPs and their bioavailability in humans is unclear. Dietary non-heme iron is physiologically absorbed via the divalent metal transporter-1 (DMT1) pathway. Using radio- iron isotope labelling in mice with a partial knockdown of intestine-specific DMT1, we assessed oral absorption and tissue biodistribution of nanostructured ferric phosphate (FePO4-NP; specific surface area [SSA] 98 m2g-1) compared to to ferrous sulfate (FeSO4), the reference compound. We show that absorption of iron from FePO4-NP appears to be largely DMT1 dependent and that its biodistribution after absorption is similar to that from FeSO4, without abnormal deposition of iron in the reticuloendothelial system. Furthermore, we demonstrate high bioavailability from iron NPs in iron deficient anemic women in a randomized, cross-over study using stable-isotope labelling: absorption and subsequent erythrocyte iron utilization from two 57Fe-labeled FePO4-NP with SSAs of 98 m2g-1 and 188 m2g-1 was 2.8-fold and 5.4-fold higher than from bulk FePO4 with an SSA of 25 m2g-1 (P < 0.001) when added to a rice and vegetable meal consumed by iron deficient anemic women. The FePO4-NP 188 m2g-1 achieved 72% relative bioavailability compared to FeSO4. These data suggest FePO4-NPs may be useful for nutritional applications.

Lack of NKG2D in MAGT1-deficient patients is caused by hypoglycosylation.

Mutations in the X-linked gene MAGT1 cause a Congenital Disorder of Glycosylation (CDG), with two distinct clinical phenotypes: a primary immunodeficiency (XMEN disorder) versus intellectual and developmental disability. It was previously established that MAGT1 deficiency abolishes steady-state expression of the immune response protein NKG2D (encoded by KLRK1) in lymphocytes. Here, we show that the reduced steady-state levels of NKG2D are caused by hypoglycosylation of the protein and we pinpoint the exact site that is underglycosylated in MAGT1-deficient patients. Furthermore, we challenge the possibility that supplementation with magnesium restores NKG2D levels and show that the addition of this ion does not significantly improve NKG2D steady-state expression nor does it rescue the hypoglycosylation defect in CRISPR-engineered human cell lines. Moreover, magnesium supplementation of an XMEN patient did not result in restoration of NKG2D expression on the cell surface of lymphocytes. In summary, we demonstrate that in MAGT1-deficient patients, the lack of NKG2D is caused by hypoglycosylation, further elucidating the pathophysiology of XMEN/MAGT1-CDG.

A Double-Blind, Placebo-Controlled, Crossover Study of Magnesium Supplementation in Patients with XMEN Disease.

X-linked MAGT1 deficiency with increased susceptibility to Epstein-Barr virus (EBV) infection and N-linked glycosylation defect (XMEN) disease is an inborn error of immunity caused by loss-of-function mutations in the magnesium transporter 1 (MAGT1) gene. The original studies of XMEN patients focused on impaired magnesium regulation, leading to decreased EBV-cytotoxicity and the loss of surface expression of the activating receptor "natural killer group 2D" (NKG2D) on CD8+ T cells and NK cells. In vitro studies showed that supraphysiological supplementation of magnesium rescued these defects. Observational studies in 2 patients suggested oral magnesium supplementation could decrease EBV viremia. Hence, we performed a randomized, double-blind, placebo-controlled, crossover study in 2 parts. In part 1, patients received either oral magnesium L-threonate (MLT) or placebo for 12 weeks followed by 12 weeks of the other treatment. Part 2 began with 3 days of high-dose intravenous (IV) magnesium sulfate (MgSO4) followed by open-label MLT for 24 weeks. One EBV-infected and 3 EBV-naïve patients completed part 1. One EBV-naïve patient was removed from part 2 of the study due to asymptomatic elevation of liver enzymes during IV MgSO4. No change in EBV or NKG2D status was observed. In vitro magnesium supplementation experiments in cells from 14 XMEN patients failed to significantly rescue NKG2D expression and the clinical trial was stopped. Although small, this study indicates magnesium supplementation is unlikely to be an effective therapeutic option in XMEN disease.

Root-to-shoot iron partitioning in Arabidopsis requires IRON-REGULATED TRANSPORTER1 (IRT1) protein but not its iron(II) transport function.

IRON-REGULATED TRANSPORTER1 (IRT1) is the root high-affinity ferrous iron (Fe) uptake system and indispensable for the completion of the life cycle of Arabidopsis thaliana without vigorous Fe supplementation. Here we provide evidence supporting a second role of IRT1 in root-to-shoot partitioning of Fe. We show that irt1 mutants overaccumulate Fe in roots, most prominently in the cortex of the differentiation zone in irt1-2, compared to the wild type. Shoots of irt1-2 are severely Fe-deficient according to Fe content and marker transcripts, as expected. We generated irt1-2 lines producing IRT1 mutant variants carrying single amino-acid substitutions of key residues in transmembrane helices IV and V, Ser206 and His232, which are required for transport activity in yeast. Root short-term 55 Fe uptake rates were uninformative concerning IRT1-mediated transport. Overall irt1-like concentrations of the secondary substrate Mn suggested that the transgenic Arabidopsis lines also remain incapable of IRT1-mediated root Fe uptake. Yet, IRT1S206A partially complements rosette dwarfing and leaf chlorosis of irt1-2, as well as root-to-shoot Fe partitioning and gene expression defects of irt1-2, all of which are fully complemented by wild-type IRT1. Taken together, these results suggest a regulatory function for IRT1 in root-to-shoot Fe partitioning that does not require Fe transport activity of IRT1. Among the genes of which transcript levels are partially dependent on IRT1, we identify MYB DOMAIN PROTEIN10, MYB DOMAIN PROTEIN72 and NICOTIANAMINE SYNTHASE4 as candidates for effecting IRT1-dependent Fe mobilization in roots. Understanding the biological functions of IRT1 will help to improve Fe nutrition and the nutritional quality of agricultural crops.

Enteral ferric citrate absorption is dependent on the iron transport protein ferroportin.

Ferric citrate is approved as an iron replacement product in patients with non-dialysis chronic kidney disease and iron deficiency anemia. Ferric citrate-delivered iron is enterally absorbed, but the specific mechanisms involved have not been evaluated, including the possibilities of conventional, transcellular ferroportin-mediated absorption and/or citrate-mediated paracellular absorption. Here, we first demonstrate the efficacy of ferric citrate in high hepcidin models, including Tmprss6 knockout mice (characterized by iron-refractory iron deficiency anemia) with and without adenine diet-induced chronic kidney disease. Next, to assess whether or not enteral ferric citrate absorption is dependent on ferroportin, we evaluated the effects of ferric citrate in a tamoxifen-inducible, enterocyte-specific ferroportin knockout murine model (Villin-Cre-ERT2, Fpnflox/flox). In this model, ferroportin deletion was efficient, as tamoxifen injection induced a 4000-fold decrease in duodenum ferroportin mRNA expression, with undetectable ferroportin protein on Western blot of duodenal enterocytes, resulting in a severe iron deficiency anemia phenotype. In ferroportin-deficient mice, three weeks of 1% ferric citrate dietary supplementation, a dose that prevented iron deficiency in control mice, did not improve iron status or rescue the iron deficiency anemia phenotype. We repeated the conditional ferroportin knockout experiment in the setting of uremia, using an adenine nephropathy model, where three weeks of 1% ferric citrate dietary supplementation again failed to improve iron status or rescue the iron deficiency anemia phenotype. Thus, our data suggest that enteral ferric citrate absorption is dependent on conventional enterocyte iron transport by ferroportin and that, in these models, significant paracellular absorption does not occur.

TMEM165 a new player in proteoglycan synthesis: loss of TMEM165 impairs elongation of chondroitin- and heparan-sulfate glycosaminoglycan chains of proteoglycans and triggers early chondrocyte differentiation and hypertrophy.

TMEM165 deficiency leads to skeletal disorder characterized by major skeletal dysplasia and pronounced dwarfism. However, the molecular mechanisms involved have not been fully understood. Here, we uncover that TMEM165 deficiency impairs the synthesis of proteoglycans by producing a blockage in the elongation of chondroitin-and heparan-sulfate glycosaminoglycan chains leading to the synthesis of proteoglycans with shorter glycosaminoglycan chains. We demonstrated that the blockage in elongation of glycosaminoglycan chains is not due to defect in the Golgi elongating enzymes but rather to availability of the co-factor Mn2+. Supplementation of cell with Mn2+ rescue the elongation process, confirming a role of TMEM165 in Mn2+ Golgi homeostasis. Additionally, we showed that TMEM165 deficiency functionally impairs TGFß and BMP signaling pathways in chondrocytes and in fibroblast cells of TMEM165 deficient patients. Finally, we found that loss of TMEM165 impairs chondrogenic differentiation by accelerating the timing of Ihh expression and promoting early chondrocyte maturation and hypertrophy. Collectively, our results indicate that TMEM165 plays an important role in proteoglycan synthesis and underline the critical role of glycosaminoglycan chains structure in the regulation of chondrogenesis. Our data also suggest that Mn2+ supplementation may be a promising therapeutic strategy in the treatment of TMEM165 deficient patients.

The Impact of ZIP8 Disease-Associated Variants G38R, C113S, G204C, and S335T on Selenium and Cadmium Accumulations: The First Characterization.

The metal cation symporter ZIP8 (SLC39A8) is a transmembrane protein that imports the essential micronutrients iron, manganese, and zinc, as well as heavy toxic metal cadmium (Cd). It has been recently suggested that selenium (Se), another essential micronutrient that has long been known for its role in human health and cancer risk, may also be transported by the ZIP8 protein. Several mutations in the ZIP8 gene are associated with the aberrant ion homeostasis of cells and can lead to human diseases. However, the intricate relationships between ZIP8 mutations, cellular Se homeostasis, and human diseases (including cancers and illnesses associated with Cd exposure) have not been explored. To further verify if ZIP8 is involved in cellular Se transportation, we first knockout (KO) the endogenous expression of ZIP8 in the HeLa cells using the CRISPR/Cas9 system. The elimination of ZIP8 expression was examined by PCR, DNA sequencing, immunoblot, and immunofluorescence analyses. Inductively coupled plasma mass spectrometry indicated that reduced uptake of Se, along with other micronutrients and Cd, was observed in the ZIP8-KO cells. In contrast, when ZIP8 was overexpressed, increased Se uptake could be detected in the ZIP8-overexpressing cells. Additionally, we found that ZIP8 with disease-associated single-point mutations G38R, G204C, and S335T, but not C113S, showed reduced Se transport ability. We then evaluated the potential of Se on Cd cytotoxicity prevention and therapy of cancers. Results indicated that Se could suppress Cd-induced cytotoxicity via decreasing the intracellular Cd transported by ZIP8, and Se exhibited excellent anticancer activity against not all but only selected cancer cell lines, under restricted experimental conditions. Moreover, clinical-based bioinformatic analyses revealed that up-regulated ZIP8 gene expression was common across multiple cancer types, and selenoproteins that were significantly co-expressed with ZIP8 in these cancers had been identified. Taken together, this study concludes that ZIP8 is an important protein in modulating cellular Se levels and provides insights into the roles of ZIP8 and Se in disease prevention and therapy.

Zinc Status Index (ZSI) for Quantification of Zinc Physiological Status.

Zinc (Zn) deficiency is estimated to affect over one billion (17%) of the world's population. Zn plays a key role in various cellular processes such as differentiation, apoptosis, and proliferation, and is used for vital biochemical and structural processes in the body. Widely used biomarkers of Zn status include plasma, whole blood, and urine Zn, which decrease in severe Zn deficiency; however, accurate assessment of Zn status, especially in mild to moderate deficiency, is difficult, as studies with these biomarkers are often contradictory and inconsistent. Thus, sensitive and specific biological markers of Zn physiological status are still needed. In this communication, we provide the Zn status index (ZSI) concept, which consists of a three-pillar formula: (1) the LA:DGLA ratio, (2) mRNA gene expression of Zn-related proteins, and (3) gut microbiome profiling to provide a clear assessment of Zn physiological status and degree of Zn deficiency with respect to assessing dietary Zn manipulation. Analysis of five selected studies found that with lower dietary Zn intake, erythrocyte LA:DGLA ratio increased, mRNA gene expression of Zn-related proteins in duodenal and liver tissues was altered, and gut microbiota populations differed, where the ZSI, a statistical model trained on data from these studies, was built to give an accurate estimation of Zn physiological status. However, the ZSI needs to be tested and refined further to determine its full potential.

Redundant roles of four ZIP family members in zinc homeostasis and seed development in Arabidopsis thaliana.

Zinc (Zn) is essential for normal plant growth and development. The Zn-regulated transporter, iron-regulated transporter (IRT)-like protein (ZIP) family members are involved in Zn transport and cellular Zn homeostasis throughout the domains of life. In this study, we have characterized four ZIP transporters from Arabidopsis thaliana (IRT3, ZIP4, ZIP6, and ZIP9) to better understand their functional roles. The four ZIP proteins can restore the growth defect of a yeast Zn uptake mutant and are upregulated under Zn deficiency. Single and double mutants show no phenotypes under Zn-sufficient or Zn-limited growth conditions. In contrast, triple and quadruple mutants show impaired growth irrespective of external Zn supply due to reduced Zn translocation from root to shoot. All four ZIP genes are highly expressed during seed development, and siliques from all single and higher-order mutants exhibited an increased number of abnormal seeds and decreased Zn levels in mature seeds relative to wild type. The seed phenotypes could be reversed by supplementing the soil with Zn. Our data demonstrate that IRT3, ZIP4, ZIP6, and ZIP9 function redundantly in maintaining Zn homeostasis and seed development in A. thaliana.

Zap70 Regulates TCR-Mediated Zip6 Activation at the Immunological Synapse.

The essential microelement zinc plays immunoregulatory roles via its ability to influence signaling pathways. Zinc deficiency impairs overall immune function and resultantly increases susceptibility to infection. Thus, zinc is considered as an immune-boosting supplement for populations with hypozincemia at high-risk for infection. Besides its role as a structural cofactor of many proteins, zinc also acts as an intracellular messenger in immune cell signaling. T-cell activation instructs zinc influx from extracellular and subcellular sources through the Zip6 and Zip8 zinc transporters, respectively. Increased cytoplasmic zinc participates in the regulation of T-cell responses by modifying activation signaling. However, the mechanism underlying the activation-dependent movement of zinc ions by Zip transporters in T cells remains elusive. Here, we demonstrate that Zip6, one of the most abundantly expressed Zip transporters in T cells, is mainly localized to lipid rafts in human T cells and is recruited into the immunological synapse in response to TCR stimulation. This was demonstrated through confocal imaging of the interaction between CD4+ T cells and antigen-presenting cells. Further, immunoprecipitation assays show that TCR triggering induces tyrosine phosphorylation of Zip6, which has at least three putative tyrosine motifs in its long cytoplasmic region, and this phosphorylation is coupled with its physical interaction with Zap70. Silencing Zip6 reduces zinc influx from extracellular sources and suppresses T-cell responses, suggesting an interaction between Zip6-mediated zinc influx and TCR activation. These results provide new insights into the mechanism through which Zip6-mediated zinc influx occurs in a TCR activation-dependent manner in human CD4+ T cells.