Biochemical characterization and subcellular localization of human copper transporter 1 (hCTR1)

Adriana E.M. KLOMP; Bastiaan B.J. TOPS; Inge E.T. vandenBERG; Ruud BERGER; Leo W.J. KLOMP

doi:10.1042/bj20011803

Biochemical characterization and subcellular localization of human copper transporter 1 (hCTR1)

Biochemical Journal ◽

10.1042/bj20011803 ◽

2002 ◽

Vol 364 (2) ◽

pp. 497-505 ◽

Cited By ~ 131

Author(s):

Adriana E.M. KLOMP ◽

Bastiaan B.J. TOPS ◽

Inge E.T. vandenBERG ◽

Ruud BERGER ◽

Leo W.J. KLOMP

Keyword(s):

Subcellular Localization ◽

Molecular Mechanisms ◽

Biochemical Characterization ◽

Copper Toxicity ◽

Menkes Disease ◽

Cell Types ◽

Polyclonal Antiserum ◽

Copper Uptake ◽

Copper Transporter ◽

Copper Transporter 1

The human copper transporter 1 gene (hCTR1) was previously identified by functional complementation in ctr1-deficient yeast. Overexpression of hCTR1 in wild-type yeast leads to increased sensitivity to copper toxicity, and mice with a homozygous disruption at the Ctr1 locus die early during embryogenesis. It is proposed that hCTR1 is responsible for high-affinity copper uptake into human cells, but the underlying molecular mechanisms are unknown. To begin to investigate the biochemical characteristics of hCTR1, a polyclonal antiserum was raised against recombinant hCTR1-fusion peptides. Biosynthetic studies using this antiserum revealed that hCTR1 was synthesized as a precursor protein of 28kDa containing N-linked oligosaccharides, and is then converted to a mature protein of approx. 35kDa, which is ubiquitously expressed. Immunofluorescence studies showed that subcellular hCTR1 localization differed markedly between cell types. In some cell lines, hCTR1 was located predominantly in an intracellular vesicular perinuclear compartment, and in others hCTR1 was located predominantly at the plasma membrane. In contrast with the copper export P-type ATPases mutated in Wilson disease and Menkes disease, the localization of hCTR1 was not influenced by copper concentrations. Inhibition of endocytosis by methyl-β-cyclodextrin caused a partial redistribution of hCTR1 to the cell surface of HeLa cells. Taken together, the results in this study suggest a cell-specific control of copper uptake, which involves subcellular localization of the hCTR1 protein.

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Characterization of the Copper Transporters from Lotus spp. and Their Involvement under Flooding Conditions

International Journal of Molecular Sciences ◽

10.3390/ijms20133136 ◽

2019 ◽

Vol 20 (13) ◽

pp. 3136 ◽

Cited By ~ 3

Author(s):

Francisco J. Escaray ◽

Cristian J. Antonelli ◽

Guillermo J. Copello ◽

Sergi Puig ◽

Lola Peñarrubia ◽

...

Keyword(s):

Molecular Mechanisms ◽

Higher Plants ◽

Economic Losses ◽

Copper Uptake ◽

Copper Transporter ◽

Copper Transporters ◽

Transporter Family ◽

Adaptive Mechanisms ◽

High Prevalence ◽

Natural Grassland

Forage legumes are an important livestock nutritional resource, which includes essential metals, such as copper. Particularly, the high prevalence of hypocuprosis causes important economic losses to Argentinian cattle agrosystems. Copper deficiency in cattle is partially due to its low content in forage produced by natural grassland, and is exacerbated by flooding conditions. Previous results indicated that incorporation of Lotus spp. into natural grassland increases forage nutritional quality, including higher copper levels. However, the biological processes and molecular mechanisms involved in copper uptake by Lotus spp. remain poorly understood. Here, we identify four genes that encode putative members of the Lotus copper transporter family, denoted COPT in higher plants. A heterologous functional complementation assay of the Saccharomyces cerevisiae ctr1∆ctr3∆ strain, which lacks the corresponding yeast copper transporters, with the putative Lotus COPT proteins shows a partial rescue of the yeast phenotypes in restrictive media. Under partial submergence conditions, the copper content of L. japonicus plants decreases and the expression of two Lotus COPT genes is induced. These results strongly suggest that the Lotus COPT proteins identified in this work function in copper uptake. In addition, the fact that environmental conditions affect the expression of certain COPT genes supports their involvement in adaptive mechanisms and envisages putative biotechnological strategies to improve cattle copper nutrition.

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Alleviation of Copper Toxicity in Arabidopsis thaliana by Silicon Addition to Hydroponic Solutions

Journal of the American Society for Horticultural Science ◽

10.21273/jashs.133.5.670 ◽

2008 ◽

Vol 133 (5) ◽

pp. 670-677 ◽

Cited By ~ 58

Author(s):

Jie Li ◽

Scott M. Leisner ◽

Jonathan Frantz

Keyword(s):

Arabidopsis Thaliana ◽

Copper Toxicity ◽

Copper Stress ◽

Copper Transporter ◽

Atpase Subunit ◽

Cu Stress ◽

Leaf Chlorosis ◽

Polymerase Chain ◽

Copper Transporter 1

Copper (Cu) is an essential micronutrient for plants and is the a.i. in pesticides for some pathogens and algae. Elevated doses of Cu can cause toxicity in plants. While silicon (Si) is reported to alleviate the toxicity of some heavy metals, its role in reducing the symptoms induced by excess Cu is unclear. Therefore, the role of Si in plant response to Cu stress was investigated in arabidopsis [Arabidopsis thaliana (L.) Heyn.]. Based on plant symptoms (a reduction of leaf chlorosis as well as increased shoot and root biomass) and a reduction of phenylalanine ammonia lyase [PAL (EC 4.3.1.5), a stress-induced enzyme] activity in the shoot, Si was found to alleviate copper stress. Real-time reverse transcriptase-polymerase chain reaction analyses indicated that the RNA levels of two arabidopsis copper transporter genes, copper transporter 1 (COPT1) and heavy metal ATPase subunit 5 (HMA5) were induced by high levels of Cu, but were significantly decreased when Si levels were also elevated. Taken together, our findings indicate that Si addition can improve the resistance of arabidopsis to Cu stress, and this improvement operates on multiple levels, ranging from physiological changes to alterations of gene expression.

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The Conformational Plasticity of the Selectivity Filter Methionines Controls the In-Cell Cu(I) Uptake through the CTR1 transporter

10.1101/2021.11.04.467269 ◽

2021 ◽

Author(s):

Pavel Janoš ◽

Jana Aupič ◽

Sharon Ruthstein ◽

Alessandra Magistrato

Keyword(s):

Cellular Uptake ◽

Selectivity Filter ◽

Copper Uptake ◽

Cellular Functions ◽

Copper Transporter ◽

High Affinity ◽

Windows Of Opportunity ◽

Conformational Plasticity ◽

Copper Transporter 1 ◽

Autoregulatory Mechanism

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 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, these outcomes resolve fundamental questions in CTR1 biology and open new windows of opportunity to tackle diseases associated with an imbalanced copper uptake.

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The N-terminal 14-mer model peptide of human Ctr1 can collect Cu(ii) from albumin. Implications for copper uptake by Ctr1

Metallomics ◽

10.1039/c8mt00274f ◽

2018 ◽

Vol 10 (12) ◽

pp. 1723-1727 ◽

Cited By ~ 15

Author(s):

Ewelina Stefaniak ◽

Dawid Płonka ◽

Simon C. Drew ◽

Karolina Bossak-Ahmad ◽

Kathryn L. Haas ◽

...

Keyword(s):

Human Serum Albumin ◽

Serum Albumin ◽

Human Serum ◽

Copper Uptake ◽

Model Peptide ◽

Copper Transporter ◽

Copper Transporter 1

The superior Cu(ii) affinity of human copper transporter 1 (hCtr1) drives copper acquisition from human serum albumin (HSA).

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Deletion of hepatic Ctr1 reveals its function in copper acquisition and compensatory mechanisms for copper homeostasis

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.90632.2008 ◽

2009 ◽

Vol 296 (2) ◽

pp. G356-G364 ◽

Cited By ~ 49

Author(s):

Heejeong Kim ◽

Hwa-Young Son ◽

Sarah M. Bailey ◽

Jaekwon Lee

Keyword(s):

Critical Role ◽

Copper Metabolism ◽

Copper Homeostasis ◽

Normal Growth ◽

Copper Uptake ◽

Compensatory Mechanisms ◽

Copper Transporter ◽

Hepatic Copper ◽

Urinary Copper ◽

Copper Transporter 1

Copper is a vital trace element required for normal growth and development of many organisms. To determine the roles for copper transporter 1 (Ctr1) in hepatic copper metabolism and the contribution of the liver to systemic copper homeostasis, we have generated and characterized mice in which Ctr1 is deleted specifically in the liver. These mice express less than 10% residual Ctr1 protein in the liver and exhibit a small but significant growth retardation, which disappears with age. Hepatic copper concentrations and the activities of copper-requiring enzymes are reduced; however, mild copper deficiency relative to Ctr1 protein deficit indicates compensatory mechanisms for copper metabolism. Copper concentrations of other organs did not alter despite the defect in hepatic copper uptake. Whereas biliary copper excretion is reduced, urinary copper concentration in these mice is higher than that of control mice. Our data indicate that Ctr1 plays a critical role in copper acquisition in the liver, and, when Ctr1 expression is compromised, compensatory mechanisms facilitate copper uptake and/or retention in the liver and excretion of copper via urine.

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Mammalian copper homeostasis requires retromer-dependent recycling of the high-affinity copper transporter 1

Journal of Cell Science ◽

10.1242/jcs.249201 ◽

2020 ◽

Vol 133 (16) ◽

pp. jcs249201 ◽

Cited By ~ 1

Author(s):

Rachel Curnock ◽

Peter J. Cullen

Keyword(s):

Cell Surface ◽

Nutrient Balance ◽

Copper Homeostasis ◽

Copper Exposure ◽

Copper Uptake ◽

Copper Transporter ◽

High Affinity ◽

Cellular Copper ◽

Copper Transporter 1 ◽

Toxicity Effects

ABSTRACTThe concentration of essential micronutrients, such as copper (used here to describe both Cu+ and Cu2+), within the cell is tightly regulated to avoid their adverse deficiency and toxicity effects. Retromer-mediated sorting and recycling of nutrient transporters within the endo-lysosomal network is an essential process in regulating nutrient balance. Cellular copper homeostasis is regulated primarily by two transporters: the copper influx transporter copper transporter 1 (CTR1; also known as SLC31A1), which controls the uptake of copper, and the copper-extruding ATPase ATP7A, a recognised retromer cargo. Here, we show that in response to fluctuating extracellular copper, retromer controls the delivery of CTR1 to the cell surface. Following copper exposure, CTR1 is endocytosed to prevent excessive copper uptake. We reveal that internalised CTR1 localises on retromer-positive endosomes and, in response to decreased extracellular copper, retromer controls the recycling of CTR1 back to the cell surface to maintain copper homeostasis. In addition to copper, CTR1 plays a central role in the trafficking of platinum. The efficacy of platinum-based cancer drugs has been correlated with CTR1 expression. Consistent with this, we demonstrate that retromer-deficient cells show reduced sensitivity to the platinum-based drug cisplatin.

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Anaphylactic Degranulation of Mast Cells: Focus on Compound Exocytosis

Journal of Immunology Research ◽

10.1155/2019/9542656 ◽

2019 ◽

Vol 2019 ◽

pp. 1-12 ◽

Cited By ~ 9

Author(s):

Ofir Klein ◽

Ronit Sagi-Eisenberg

Keyword(s):

Mast Cell ◽

Mast Cells ◽

Molecular Mechanisms ◽

Secretory Granules ◽

Cell Types ◽

Secretory Cells ◽

Regulated Exocytosis ◽

Open Questions ◽

Conflicting Evidence ◽

Compound Exocytosis

Anaphylaxis is a notorious type 2 immune response which may result in a systemic response and lead to death. A precondition for the unfolding of the anaphylactic shock is the secretion of inflammatory mediators from mast cells in response to an allergen, mostly through activation of the cells via the IgE-dependent pathway. While mast cells are specialized secretory cells that can secrete through a variety of exocytic modes, the most predominant mode exerted by the mast cell during anaphylaxis is compound exocytosis—a specialized form of regulated exocytosis where secretory granules fuse to one another. Here, we review the modes of regulated exocytosis in the mast cell and focus on compound exocytosis. We review historical landmarks in the research of compound exocytosis in mast cells and the methods available for investigating compound exocytosis. We also review the molecular mechanisms reported to underlie compound exocytosis in mast cells and expand further with reviewing key findings from other cell types. Finally, we discuss the possible reasons for the mast cell to utilize compound exocytosis during anaphylaxis, the conflicting evidence in different mast cell models, and the open questions in the field which remain to be answered.

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Genetic analysis of amyotrophic lateral sclerosis identifies contributing pathways and cell types

Science Advances ◽

10.1126/sciadv.abd9036 ◽

2021 ◽

Vol 7 (3) ◽

pp. eabd9036

Author(s):

Sara Saez-Atienzar ◽

Sara Bandres-Ciga ◽

Rebekah G. Langston ◽

Jonggeol J. Kim ◽

Shing Wan Choi ◽

...

Keyword(s):

Amyotrophic Lateral Sclerosis ◽

Membrane Trafficking ◽

Molecular Mechanisms ◽

Cell Types ◽

Polygenic Risk Score ◽

Genome Wide ◽

Genome Wide Data ◽

Data Driven Approach ◽

Single Nucleus ◽

Lateral Sclerosis

Despite the considerable progress in unraveling the genetic causes of amyotrophic lateral sclerosis (ALS), we do not fully understand the molecular mechanisms underlying the disease. We analyzed genome-wide data involving 78,500 individuals using a polygenic risk score approach to identify the biological pathways and cell types involved in ALS. This data-driven approach identified multiple aspects of the biology underlying the disease that resolved into broader themes, namely, neuron projection morphogenesis, membrane trafficking, and signal transduction mediated by ribonucleotides. We also found that genomic risk in ALS maps consistently to GABAergic interneurons and oligodendrocytes, as confirmed in human single-nucleus RNA-seq data. Using two-sample Mendelian randomization, we nominated six differentially expressed genes (ATG16L2, ACSL5, MAP1LC3A, MAPKAPK3, PLXNB2, and SCFD1) within the significant pathways as relevant to ALS. We conclude that the disparate genetic etiologies of this fatal neurological disease converge on a smaller number of final common pathways and cell types.

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In-depth transcriptomic analysis of human retina reveals molecular mechanisms underlying diabetic retinopathy

Scientific Reports ◽

10.1038/s41598-021-88698-3 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Kolja Becker ◽

Holger Klein ◽

Eric Simon ◽

Coralie Viollet ◽

Christian Haslinger ◽

...

Keyword(s):

Diabetic Retinopathy ◽

Rna Sequencing ◽

Molecular Mechanisms ◽

Vision Loss ◽

Ganglion Cells ◽

Expression Profiles ◽

Cell Types ◽

Sequencing Data ◽

Disease Stages ◽

Post Transcriptional Regulation

AbstractDiabetic Retinopathy (DR) is among the major global causes for vision loss. With the rise in diabetes prevalence, an increase in DR incidence is expected. Current understanding of both the molecular etiology and pathways involved in the initiation and progression of DR is limited. Via RNA-Sequencing, we analyzed mRNA and miRNA expression profiles of 80 human post-mortem retinal samples from 43 patients diagnosed with various stages of DR. We found differentially expressed transcripts to be predominantly associated with late stage DR and pathways such as hippo and gap junction signaling. A multivariate regression model identified transcripts with progressive changes throughout disease stages, which in turn displayed significant overlap with sphingolipid and cGMP–PKG signaling. Combined analysis of miRNA and mRNA expression further uncovered disease-relevant miRNA/mRNA associations as potential mechanisms of post-transcriptional regulation. Finally, integrating human retinal single cell RNA-Sequencing data revealed a continuous loss of retinal ganglion cells, and Müller cell mediated changes in histidine and β-alanine signaling. While previously considered primarily a vascular disease, attention in DR has shifted to additional mechanisms and cell-types. Our findings offer an unprecedented and unbiased insight into molecular pathways and cell-specific changes in the development of DR, and provide potential avenues for future therapeutic intervention.

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