High-affinity Copper Transporter Research Articles

Copper Dysmetabolism is Connected to Epithelial-Mesenchymal Transition: A Pilot Study in Colorectal Cancer Patients.

Colorectal cancer (CRC) is among the most diagnosed cancers worldwide, whose risk of mortality is associated with the development of metastases to the liver, lungs, and peritoneum. Of note, CRC is highly dependent on copper to sustain its proliferation and aggressiveness. Copper acts not only as a pivotal cofactor for several cuproproteins but also as an allosteric modulator of kinases essential to fulfill the epithelial-to-mesenchymal-transition (EMT), the main mechanism driving cancer cell spreading. System biology identified the APP and SOD1 genes among the top 10 genes shared between CRC and copper metabolism, as confirmed by the upregulation of the protein/mRNA levels of APP observed in CRC tissues. The significant increase of copper found in the sera of CRC patients was paralleled by a strong reduction of copper in the CRC tissues, in agreement with the decreased level of the high-affinity copper transporter CTR1 mRNA (SLC31A1) and LOXL2. As expected, in CRC tissues the mesenchymal marker fibronectin was significantly increased, whereas vimentin and vinculin protein levels were decreased compared to adjacent healthy mucosa. Interestingly, correlation analysis showed an interconnection between vinculin and both CCS and APP. A positive correlation was also observed between APP mRNA and both CDH1 and SOD1 mRNAs. Overall, we demonstrate a correlation between cell copper imbalance and CRC progression via EMT. The results obtained lay the scientific basis for further investigation to describe the kinetics of copper dysregulation during CRC progression and to identify the main cuproproteins involved in the modulation of EMT.

Characterization of a High-Affinity Copper Transporter CTR1a in the White-Nose Syndrome Causing Fungal Pathogen Pseudogymnoascus destructans.

Copper is an essential micronutrient and the ability to scavenge tightly bound or trace levels of copper ions at the host-pathogen interface is vital for fungal proliferation in animal hosts. Recent studies suggest that trace metal ion acquisition is critical for the establishment and propagation of Pseudogymnoascus destructans, the fungal pathogen responsible for white-nose syndrome (WNS), on their bat host. However, little is known about these metal acquisition pathways in P. destructans. In this study, we report the characterization of the P. destructans high-affinity copper transporter VC83_00191 (PdCTR1a), which is implicated as a virulence factor associated with the WNS disease state. Using Saccharomyces cerevisiae as a recombinant expression host, we find that PdCTR1a can efficiently traffic Cu ions into the yeast cytoplasm. Complementary studies in the native P. destructans fungus provide evidence that PdCTR1a transcripts and protein levels are dictated by Cu-bioavailability in the growth media. Our study demonstrates that PdCTR1a is a functional high-affinity copper transporter and is relevant to Cu homeostasis pathways in P. destructans.

Characterization of a High-Affinity Copper Transporter in the White-Nose Syndrome Causing Fungal Pathogen Pseudogymnoascus destructans.

Copper is an essential micronutrient and the ability to scavenge tightly bound or trace levels of copper ions at the host-pathogen interface is vital for fungal proliferation in animal hosts. Recent studies suggest that trace metal ion acquisition is critical for the establishment and propagation of Pseudogymnoascus destructans, the fungal pathogen responsible for white-nose syndrome (WNS), on their bat host. However, little is known about these metal acquisition pathways in P. destructans. In this study, we report the characterization of the P. destructans high-affinity copper transporter VC83_00191 (PdCTR1a), which is implicated as a virulence factor associated with the WNS disease state. Using Saccharomyces cerevisiae as a recombinant expression host, we find that PdCTR1a localizes to the cell surface plasma membrane and can efficiently traffic Cu-ions into the yeast cytoplasm. Complementary studies in the native P. destructans fungus provide evidence that PdCTR1a transcripts and protein levels are dictated by Cu-bioavailability in the growth media. Our study demonstrates that PdCTR1a is a functional high-affinity copper transporter and is relevant to Cu-homeostasis pathways in P. destructans.

Role of SLC31A1 in prognosis and immune infiltration in breast cancer: a novel insight.

Copper, an essential metal element for humans, plays vital functions in cancer prognosis and immunity. SLC31A1, a high-affinity copper transporter, helps regulate copper homeostasis and has been implicated in tumor prognosis through mechanisms such as drug resistance, autophagy, ferroptosis, and cuproptosis. However, the role of SLC31A1 in breast cancer (BRCA) and its association with tumor immune infiltration has not been fully elucidated. This study aimed to investigate the expression pattern of SLC31A1, its clinical significance, and its effect on tumor immune infiltration in BRCA. We comprehensively analyzed multiple datasets, including Gene Expression Profiling Interaction Analysis (GEPIA), Tumor Immune Estimation Resource (TIMER), UALCAN, and Kaplan-Meier (KM) plotter, to assess the expression of SLC31A1 and its prognostic value in BRCA. Additionally, TIMER and TISIDB were used to explore the correlation between SLC31A1 expression and the extent of tumor immune infiltration. SLC31A1 was significantly upregulated in BRCA tissues compared to adjacent non-tumor tissues. Higher SLC31A1 expression levels were associated with poorer clinical outcome. Multivariate Cox regression analysis confirmed that SLC31A1 served as an independent prognostic indicator. Furthermore, SLC31A1 expression showed significant associations with various immunomodulators, chemokines, chemokine receptors, and tumor-infiltrating lymphocytes (TILs), including CD8+ T cells, CD4+ T cells, regulatory T cells (Tregs), follicular helper T cells (Tfh), neutrophils, M2 macrophages, tumor-associated macrophages (TAMs), and monocytes. These findings suggest that SLC31A1 may regulate macrophage polarization and T cell exhaustion in BRCA, contributing to immune evasion and poor prognosis. Our study underscores the importance of further research to explore the therapeutic potential of targeting SLC31A1 and to uncover its additional roles in BRCA beyond the known mechanisms of drug resistance, autophagy, ferroptosis, and cuproptosis.

Arabidopsis COPT1 copper transporter uses a single histidine to regulate transport activity and protein stability

Copper acquisition and subsequent delivery to target proteins are essential for many biological processes. However, the cellular levels of this trace element must be controlled because of its potential toxicity. The COPT1 protein rich in potential metal-binding amino acids functions in high affinity copper uptake at the plasma membrane of Arabidopsis cells. The functional role of these putative metal-binding residues is largely unknown. Through truncations and site-directed mutagenesis, we identified His43, a single residue within the extracellular N-terminal domain as absolutely critical for copper uptake of COPT1. Substitution of this residue with leucine, methionine or cysteine almost inactivated transport function of COPT1, implying that His43 fails to serves as a copper ligand in the regulation of COPT1 activity. Deletion of all extracellular N-terminal metal-binding residues completely blocked copper-stimulated degradation but did not alter the subcellular distribution and multimerization of COPT1. Although mutation of His43 to alanine and serine retained the transporter activity in yeast cells, the mutant protein was unstable and degraded in the proteasome in Arabidopsis cells. Our results demonstrate a pivotal role for the extracellular residue His43 in high affinity copper transport activity, and suggest common molecular mechanisms for regulating both metal transport and protein stability of COPT1.

Properties of recombinant extracellular N-terminal domain of human high-affinity copper transporter 1 (hNdCTR1) and its interactions with Cu(II) and Ag(I) ions.

High-affinity copper transporter 1 (CTR1) is a key link in the transfer of copper (Cu) from the extracellular environment to the cell. Violation in the control system of its expression, or mutations in this gene, cause a global copper imbalance. However, the mechanism of copper transfer via CTR1 remains unclear. It has been shown that transformed bacteria synthesizing the fused GB1-NdCTR become resistant to toxic silver ions. According to UV-Vis spectrophotometry and isothermal titration calorimetry, electrophoretically pure GB1-NdCTR specifically and reversibly binds copper and silver ions, and binding is associated with aggregation. Purified NdCTR1 forms SDS-resistant oligomers. The link between nontrivial properties of NdCTR1 and copper import mechanism from extracellular space, as well as potential chelating properties of NdCTR1, are discussed.

Antifungal Effect of Copper Nanoparticles against Fusarium kuroshium, an Obligate Symbiont of Euwallacea kuroshio Ambrosia Beetle.

Copper nanoparticles (Cu-NPs) have shown great antifungal activity against phytopathogenic fungi, making them a promising and affordable alternative to conventional fungicides. In this study, we evaluated the antifungal activity of Cu-NPs against Fusarium kuroshium, the causal agent of Fusarium dieback, and this might be the first study to do so. The Cu-NPs (at different concentrations) inhibited more than 80% of F. kuroshium growth and were even more efficient than a commercial fungicide used as a positive control (cupric hydroxide). Electron microscopy studies revealed dramatic damage caused by Cu-NPs, mainly in the hyphae surface and in the characteristic form of macroconidia. This damage was visible only 3 days post inoculation with used treatments. At a molecular level, the RNA-seq study suggested that this growth inhibition and colony morphology changes are a result of a reduced ergosterol biosynthesis caused by free cytosolic copper ions. Furthermore, transcriptional responses also revealed that the low- and high-affinity copper transporter modulation and the endosomal sorting complex required for transport (ESCRT) are only a few of the distinct detoxification mechanisms that, in its conjunction, F. kuroshium uses to counteract the toxicity caused by the reduced copper ion.

OP004: Newly identified disorder of copper metabolism caused by variants in CTR1, a high-affinity copper transporter

Copper is a micronutrient essential for numerous cellular functions including processes mediated by a network of copper-dependent enzymes that exploit this metal’s facility for electron exchange. The high-affinity copper transporter CTR1 is encoded by CTR1 (SLC31A1), a gene locus for which no detailed genotype-phenotype correlations have been reported. We describe identical twin boys homozygous for a novel missense pathogenic variant NM_001859.4:c.284 G>A (p.Arg95His) in the SLC31A1 gene with a distinctive autosomal recessive syndrome of infantile seizures and neurodegeneration, consistent with profound central nervous system copper deficiency.

Dynamical interplay between the human high-affinity copper transporter hCtr1 and its cognate metal ion

Abnormal cellular copper levels have been clearly implicated in genetic diseases, cancer, and neurodegeneration. Ctr1, a high-affinity copper transporter, is a homotrimeric integral membrane protein that provides the main route for cellular copper uptake. Together with a sophisticated copper transport system, Ctr1 regulates Cu(I) metabolism in eukaryotes. Despite its pivotal role in normal cell function, the molecular mechanism of copper uptake and transport via Ctr1 remains elusive. In this study, electron paramagnetic resonance (EPR), UV-visible spectroscopy, and all-atom simulations were employed to explore Cu(I) binding to full-length human Ctr1 (hCtr1), thereby elucidating how metal binding at multiple distinct sites affects the hCtr1 conformational dynamics. We demonstrate that each hCtr1 monomer binds up to five Cu(I) ions and that progressive Cu(I) binding triggers a marked structural rearrangement in the hCtr1 C-terminal region. The observed Cu(I)-induced conformational remodeling suggests that the C-terminal region may play a dual role, serving both as a channel gate and as a shuttle mediating the delivery of copper ions from the extracellular hCtr1 selectivity filter to intracellular metallochaperones. Our findings thus contribute to a more complete understanding of the mechanism of hCtr1-mediated Cu(I) uptake and provide a conceptual basis for developing mechanism-based therapeutics for treating pathological conditions linked to de-regulated copper metabolism.

The conformational plasticity of the selectivity filter methionines controls the in-cell Cu(I) uptake through the CTR1 transporter.

Copper is a trace element vital to many cellular functions. Yet its abnormal levels are toxic to cells, provoking a variety of severe diseases. The high affinity copper transporter 1 (CTR1), being the main in-cell copper [Cu(I)] entry route, tightly regulates its cellular uptake via a still elusive mechanism. Here, all-atoms simulations unlock the molecular terms of Cu(I) transport in eukaryotes disclosing that the two methionine (Met) triads, forming the selectivity filter, play an unprecedented dual role both enabling selective Cu(I) transport and regulating its uptake rate thanks to an intimate coupling between the conformational plasticity of their bulky side chains and the number of bound Cu(I) ions. Namely, the Met residues act as a gate reducing the Cu(I) import rate when two ions simultaneously bind to CTR1. This may represent an elegant autoregulatory mechanism through which CTR1 protects the cells from excessively high, and hence toxic, in-cell Cu(I) levels. Overall, our outcomes resolve fundamental questions in CTR1 biology and open new windows of opportunity to tackle diseases associated with an imbalanced copper uptake.

Identification and molecular characterization of the high-affinity copper transporters family in Solanum lycopersicum

Copper (Cu) plays a key role as cofactor in the plant proteins participating in essential cellular processes, such as electron transport and free radical scavenging. Despite high-affinity Cu transporters (COPTs) being key participants in Cu homeostasis maintenance, very little is known about COPTs in tomato (Solanum lycopersicum) even though it is the most consumed fruit worldwide and this crop is susceptible to suboptimal Cu conditions. In this study, a six-member family of COPT (SlCOPT1-6) was identified and characterized. SlCOPTs have a conserved architecture consisting of three transmembrane domains and β-strains. However, the presence of essential methionine residues, a methionine-enriched amino-terminal region, an Mx3Mx12Gx3G Cu-binding motif and a cysteine rich carboxy-terminal region, all required for their functionality, is more variable among members. Accordingly, functional complementation assays in yeast indicate that SlCOPT1 and SlCOPT2 are able to transport Cu inside the cell, while SlCOPT3 and SlCOPT5 are only partially functional. In addition, protein interaction network analyses reveal the connection between SlCOPTs and Cu PIB-type ATPases, other metal transporters, and proteins related to the peroxisome. Gene expression analyses uncover organ-dependency, fruit vasculature tissue specialization and ripening-dependent gene expression profiles, as well as different response to Cu deficiency or toxicity in an organ-dependent manner.

Targeting the Copper Transport System to Improve Treatment Efficacies of Platinum-Containing Drugs in Cancer Chemotherapy.

The platinum (Pt)-containing antitumor drugs including cisplatin (cis-diamminedichloroplatinum II, cDDP), carboplatin, and oxaliplatin, have been the mainstay of cancer chemotherapy. These drugs are effective in treating many human malignancies. The major cell-killing target of Pt drugs is DNA. Recent findings underscored the important roles of Pt drug transport system in cancer therapy. While many mechanisms have been proposed for Pt-drug transport, the high-affinity copper transporter (hCtr1), Cu chaperone (Atox1), and Cu exporters (ATP7A and ATP7B) are also involved in cDDP transport, highlighting Cu homeostasis regulation in Pt-based cancer therapy. It was demonstrated that by reducing cellular Cu bioavailable levels by Cu chelators, hCtr1 is transcriptionally upregulated by transcription factor Sp1, which binds the promoters of Sp1 and hCtr1. In contrast, elevated Cu poisons Sp1, resulting in suppression of hCtr1 and Sp1, constituting the Cu-Sp1-hCtr1 mutually regulatory loop. Clinical investigations using copper chelator (trientine) in carboplatin treatment have been conducted for overcoming Pt drug resistance due in part to defective transport. While results are encouraging, future development may include targeting multiple steps in Cu transport system for improving the efficacies of Pt-based cancer chemotherapy. The focus of this review is to delineate the mechanistic interrelationships between Cu homeostasis regulation and antitumor efficacy of Pt drugs.

Ubiquitin-independent proteasome system is required for degradation of Arabidopsis COPPER TRANSPORTER 2

Plants have evolved sophisticated mechanisms to adjust to deficiency or excess of nutrients. Membrane transport proteins play a central role in nutrient uptake from soil. In Arabidopsis thaliana, the COPPER TRANSPORTOR (COPT) family encodes high-affinity copper transporters. COPT2 is transcriptionally regulated in response to changing levels of cellular copper. However, little is known about whether COPT2 activity is subject to multiple levels of regulation. Here, we showed that the plasma membrane-/endoplasmic reticulum-resident COPT2 protein is degraded in response to excess copper. Confocal microscopy analysis together with pharmacological treatment with a vesicle trafficking inhibitor or vacuolar ATPase inhibitor indicated that copper-mediated downregulation of COPT2 is unlikely to be controlled by endosomal recycling and vacuolar system. However, COPT2 protein is stabilized by proteasome inhibition. Through site-directed mutagenesis, we found that COPT2 cannot be ubiquitinated, and lysine residues at the C-terminus is dispensable for copper-induced degradation of COPT2 but required for copper acquisition. Altogether, our findings reveal that unlike many metal transporters in Arabidopsis, COPT2 is a substrate of ubiquitin-independent proteasomal degradation but not of vacuolar proteases. These findings highlight the mechanistic diversity and complexity of plasma membrane transporter degradation.

Copper Homeostasis in Aspergillus nidulans Involves Coordinated Transporter Function, Expression and Cellular Dynamics.

Copper ion homeostasis involves a finely tuned and complex multi-level response system. This study expands on various aspects of the system in the model filamentous fungus Aspergillus nidulans. An RNA-seq screen in standard growth and copper toxicity conditions revealed expression changes in key copper response elements, providing an insight into their coordinated functions. The same study allowed for the deeper characterization of the two high-affinity copper transporters: AnCtrA and AnCtrC. In mild copper deficiency conditions, the null mutant of AnctrC resulted in secondary level copper limitation effects, while deletion of AnctrA resulted in primary level copper limitation effects under extreme copper scarcity conditions. Each transporter followed a characteristic expression and cellular localization pattern. Although both proteins partially localized at the plasma membrane, AnCtrC was visible at membranes that resembled the ER, whilst a substantial pool of AnCtrA accumulated in vesicular structures resembling endosomes. Altogether, our results support the view that AnCtrC plays a major role in covering the nutritional copper requirements and AnCtrA acts as a specific transporter for extreme copper deficiency scenarios.

Arabidopsis COPPER TRANSPORTER 1 undergoes degradation in a proteasome-dependent manner.

The essential nutrient copper is toxic in excess. Therefore, plants must tightly control copper uptake and distribution. Arabidopsis thaliana high-affinity copper transporters (COPTs) mediate copper uptake, partitioning, and redistribution. Here we show that COPT1 localizes to the plasma membrane and endoplasmic reticulum in stably transgenic plants expressing a COPT1-green fluorescent protein (GFP) fusion protein, and the fusion protein is rapidly degraded upon plant exposure to excess copper. MG132 treatment largely abolished copper-induced degradation of COPT1, implying a link between the proteasome and COPT1 activity in modulating copper uptake. Co-immunoprecipitation analyses revealed that COPT1 cannot be ubiquitinated in the presence of excess copper and MG132. Through site-directed mutagenesis, we identified Lys159 in the C-terminal cytoplasmic tail of COPT1 as critical for copper acquisition, but not for copper-mediated down-regulation of COPT1, in plants. Furthermore, pharmacological analysis showed that treatment with a vesicle trafficking inhibitor or a V-ATPase inhibitor does not alter the subcellular dynamics of COPT1-GFP, consistent with the absence of a connection between the endosomal recycling/vacuolar system and COPT1 degradation. Together, our data suggest that proteasomal degradation rather than vacuolar proteolysis is important for the regulation of copper transport to maintain copper homeostasis in plants.

Deregulated High Affinity Copper Transport Alters Iron Homeostasis in Arabidopsis.

The present work describes the effects on iron homeostasis when copper transport was deregulated in Arabidopsis thaliana by overexpressing high affinity copper transporters COPT1 and COPT3 (COPTOE). A genome-wide analysis conducted on COPT1OE plants, highlighted that iron homeostasis gene expression was affected under both copper deficiency and excess. Among the altered genes were those encoding the iron uptake machinery and their transcriptional regulators. Subsequently, COPTOE seedlings contained less iron and were more sensitive than controls to iron deficiency. The deregulation of copper (I) uptake hindered the transcriptional activation of the subgroup Ib of basic helix-loop-helix (bHLH-Ib) factors under copper deficiency. Oppositely, copper excess inhibited the expression of the master regulator FIT but activated bHLH-Ib expression in COPTOE plants, in both cases leading to the lack of an adequate iron uptake response. As copper increased in the media, iron (III) was accumulated in roots, and the ratio iron (III)/iron (II) was increased in COPTOE plants. Thus, iron (III) overloading in COPTOE roots inhibited local iron deficiency responses, aimed to metal uptake from soil, leading to a general lower iron content in the COPTOE seedlings. These results emphasized the importance of appropriate spatiotemporal copper uptake for iron homeostasis under non-optimal copper supply. The understanding of the role of copper uptake in iron metabolism could be applied for increasing crops resistance to iron deficiency.

Antagonistic bioaccumulation of waterborne Cu(II) and Cd(II) in common carp (Cyprinus carpio) and effects on ion-homeostasis and defensive mechanisms

In the aquatic environment, metals are present as mixtures, therefore studies on mixture toxicity are crucial to thoroughly understand their toxic effects on aquatic organisms. Common carp (Cyprinus carpio) were used to assess the effects of short-term Cu(II) and Cd(II) mixtures, using a fixed concentration of one of the metals, representing 25 % of its individual 96h-LC50 (concentration lethal for 50 % of the population) combined with a variable concentration of the other metal corresponding to 10, 25 or 50 % of its 96h-LC50, and vice versa. Our results showed a fast Cu and Cd bioaccumulation, with the percentage of increase in the order gill > liver > carcass. An inhibitory effect of Cu on Cd uptake was observed; higher Cu concentrations at fixed Cd levels resulted in a decreased accumulation of Cd. The presence of the two metal ions resulted in losses of total Na, K and Ca. Fish tried to compensate for the Na loss through the induction of the genes coding for Na+/K+-ATPase and H+-ATPase. Additionally, a counterintuitive induction of the gene encoding the high affinity copper transporter (CTR1) occurred, while a downregulation was expected to prevent further metal ion uptake. An induction of defensive mechanisms, both metal ion binding protein and anti-oxidant defences, was observed. Despite the metal accumulation and electrolyte loss, the low mortality suggest that common carp is able to cope with these metal levels, at least during a one-week exposure.

Functional analyses of copper transporter genes in the human liver cell line HepG2

Copper (Cu) is an essential element regulated by four genes (hCTR1, hATOX1, hATP7A, and hATP7B in humans and zctr1, zatox1, zatp7a, and zatp7b in zebrafish) in copper uptake, distribution, and transport in animal cells. Zebrafish (Danio rerio) shows a higher endogenous ratio of zatp7a to zatp7b in the liver, is relatively intolerant to copper ions and has a different zatp7a and zatp7b expression patterns in different organs. As high-affinity copper transporters, both zctr1 and hCTR1 increased copper toxicity, whereas hATOX1 and zatox1 slightly reduced copper toxicity in HepG2 cells after copper administration for 24 h. The transfected zatp7b functioned in HepG2 cells as effectively as hATP7B after both 24-h and 96-h copper exposure, but zatp7a failed to function in HepG2 cells as effectively as hATP7A. Our findings suggest that ATP7A dysfunction would increase cytotoxicity in the liver; the reason for zebrafish's copper intolerance could be the bulk dysfunction and abnormal localization of zATP7A.

CRISP-R/Cas9 Mediated Deletion of Copper Transport Genes CTR1 and DMT1 in NSCLC Cell Line H1299. Biological and Pharmacological Consequences.

Copper, the highly toxic micronutrient, plays two essential roles: it is a catalytic and structural cofactor for Cu-dependent enzymes, and it acts as a secondary messenger. In the cells, copper is imported by CTR1 (high-affinity copper transporter 1), a transmembrane high-affinity copper importer, and DMT1 (divalent metal transporter). In cytosol, enzyme-specific chaperones receive copper from CTR1 C-terminus and deliver it to their apoenzymes. DMT1 cannot be a donor of catalytic copper because it does not have a cytosol domain which is required for copper transfer to the Cu-chaperons that assist the formation of cuproenzymes. Here, we assume that DMT1 can mediate copper way required for a regulatory copper pool. To verify this hypothesis, we used CRISPR/Cas9 to generate H1299 cell line with CTR1 or DMT1 single knockout (KO) and CTR1/DMT1 double knockout (DKO). To confirm KOs of the genes qRT-PCR were used. Two independent clones for each gene were selected for further studies. In CTR1 KO cells, expression of the DMT1 gene was significantly increased and vice versa. In subcellular compartments of the derived cells, copper concentration dropped, however, in nuclei basal level of copper did not change dramatically. CTR1 KO cells, but not DMT1 KO, demonstrated reduced sensitivity to cisplatin and silver ions, the agents that enter the cell through CTR1. Using single CTR1 and DMT1 KO, we were able to show that both, CTR1 and DMT1, provided the formation of vital intracellular cuproenzymes (SOD1, COX), but not secretory ceruloplasmin. The loss of CTR1 resulted in a decrease in the level of COMMD1, XIAP, and NF-κB. Differently, the DMT1 deficiency induced increase of the COMMD1, HIF1α, and XIAP levels. The possibility of using CTR1 KO and DMT1 KO cells to study homeodynamics of catalytic and signaling copper selectively is discussed.

Copper transporter COPT5 participates in the crosstalk between vacuolar copper and iron pools mobilisation

Copper (Cu) deficiency affects iron (Fe) homeostasis in several plant processes, including the increased Fe requirements due to cuproprotein substitutions for the corresponding Fe counterpart. Loss-of-function mutants from Arabidopsis thaliana high affinity copper transporter COPT5 and Fe transporters NATURAL RESISTANCE-ASSOCIATED MACROPHAGE PROTEIN 3/4 (NRAMP3 and NRAMP4) were used to study the interaction between metals internal pools. A physiological characterisation showed that the copt5 mutant is sensitive to Fe deficiency, and that nramp3nramp4 mutant growth was severely affected under limiting Cu. By a transcriptomic analysis, we observed that NRAMP4 expression was highly induced in the copt5 mutant under Cu deficiency, while COPT5 was overexpressed in the nramp3nramp4 mutant. As a result, an enhanced mobilisation of the vacuolar Cu or Fe pools, when the other metal export through the tonoplast is impaired in the mutants, has been postulated. However, metals coming from internal pools are not used to accomplish the increased requirements that derive from metalloprotein substitution under metal deficiencies. Instead, the metal concentrations present in aerial parts of the copt5 and nramp3nramp4 mutants conversely show compensated levels of these two metals. Together, our data uncover an interconnection between Cu and Fe vacuolar pools, whose aim is to fulfil interorgan metal translocation.