Cancer remains the second leading cause of death worldwide, emphasizing the critical need for effective treatment and control strategies. Essential minerals such as copper, iron, zinc, selenium, phosphorous, calcium, and magnesium are integral to various biological processes and significantly influence cancer progression through altered metabolic pathways. For example, dysregulated copper levels promote tumor growth, while cancer cells exhibit an increased dependency on iron for signaling and redox reactions. Zinc influences tumor development through pathways such as Akt-p21. Selenium, primarily through its role in selenoproteins, exhibits anticancer potential but may also contribute to tumor progression. Similarly, dietary phosphate exacerbates tumorigenesis, metastasis, and angiogenesis through signaling pathway activation. Calcium, the most abundant mineral in the body, is tightly regulated within cells, and its dysregulation is a hallmark of various cancers. Magnesium deficiency, on the other hand, promotes cancer progression by fostering inflammation and free radical-induced DNA mutations. Interestingly, magnesium also plays a dual role, with low levels enhancing epithelial-mesenchymal transition (EMT), a critical process in cancer metastasis. This complex interplay of essential minerals underscores their potential as therapeutic targets. Dysregulation of these minerals and their pathways could be exploited to selectively target cancer cells, offering novel therapeutic strategies. This review summarizes current research on the abnormal accumulation or depletion of these microelements in tumor biology, drawing evidence from animal models, cell lines, and clinical samples. We also highlight the potential of these minerals as biomarkers for cancer diagnosis and prognosis, as well as therapeutic approaches involving metal chelators, pharmacological agents, and nanotechnology. By highlighting the intricate roles of these minerals in cancer biology, we aim to inspire further research in this critical yet underexplored area of oncology.

Food security is defined as uninterrupted access to food that meets people's dietary needs. One essential trace element of a complete diet is zinc, which is vital for various processes, including growth, development, and the immune response. The estimated global prevalence of zinc deficiency is around 30%. Meat and meat products provide an abundant and also bioavailable source of zinc. However, in developing countries, access to meat is restricted, and in developed countries, meat consumption has declined for ethical and environmental reasons. The potential for zinc deficiency arises from (i) low concentrations of this element in plant-based diets, (ii) poor zinc absorption from plant-based food in the human intestine, and (iii) the risk of uptake of toxic metals together with essential ones. This review summarises the current knowledge concerning type 4 metallothioneins, which represent promising targets for zinc biofortification. We describe their place in the zinc route from soil to seed, their expression patterns, their role in plants, and their three-dimensional protein structure and how this affects their selectivity towards zinc. This review aims to provide a comprehensive theoretical basis for the potential use of type 4 plant metallothioneins to create zinc-biofortified crops.

This study examined the correlation between multi-metal exposure and bone mineral density (BMD) in U.S. children and adolescents.

Data from 1,591 participants (aged 8-19) were analyzed using the National Health and Nutrition Examination Survey (NHANES) 2011-2016. We measured serum copper (Cu), selenium (Se), zinc (Zn), and blood lead (Pb), cadmium (Cd), mercury (Hg), manganese (Mn). Dual-energy X-ray absorptiometry assessed lumbar and total BMD. Advanced statistical approaches including weighted quantile sum (WQS) regression and bayesian kernel machine regression (BKMR) were employed to evaluate complex exposure interactions.

Blood Pb and serum Cu showed inverse associations with, while serum Se positively correlated with lumbar BMD (blood Pb: β: -0.013, serum Cu: β: -0.063, serum Se: 0.035) (all P < 0.05). The WQS index showed a significant association with both lumbar BMD(β = 0.019, P < 0.05) and total BMD (β = 0.019, P < 0.001). WQS analysis identified Cd, Se, and Hg as primary contributors to both lumbar and total BMD variations. BKMR models revealed nonlinear exposure-response relationships and synergistic effects between Cd and Mn.

These findings highlight the importance of considering mixed metal exposures in bone health assessments, providing crucial insights for developing preventive strategies to protect skeletal development in pediatric populations.

Metallothioneins (MTs) are small cysteine-rich proteins that preferentially bind d10 metal ions such as ZnII, CuI, and CdII, playing essential roles in metal ion homeostasis and detoxification. The Ec-1 metallothionein from Triticum aestivum (common bread wheat) was the first plant metallothionein for which a 3D structure was successfully determined, although this structure represents only the fully metalated state of the protein. In this study, we aim to elucidate the metalation pathway of the βE-domain of wheat Ec-1. This domain features a mononuclear ZnII binding site composed of two cysteine and two highly conserved histidine residues, reminiscent of the Zn-finger motifs found in certain proteins. Moreover, the domain forms a trinuclear Zn3Cys9 cluster, similar to the β-cluster motif observed, for example, in vertebrate MTs. To investigate the metalation pathway of the βE-domain, we combined nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry, and targeted cysteine modification techniques. Our results confidently identify the sequential binding site regions for each of the four ZnII ions and reveal intriguing, unexpected insights into the folding pathway of the peptide backbone.

Abdominal aortic aneurysm (AAA) is a life-threatening vascular condition. Currently, there are no clinically available pharmacological interventions that can stop the progression of AAA, primarily due to the incomplete understanding of its pathogenesis and the absence of effective drug delivery systems. The present study aimed to develop a targeted therapy for AAA through a nanomedicine approach involving site-specific regulation of neutrophil extracellular trap (NET)-related pathological vascular remodeling. We found that metallothionein 1 (MT1) was upregulated in AAA lesions in both humans and mice. MT1 also facilitated the formation of NETs and subsequently induced phenotypic transformation and apoptosis in vascular smooth muscle cells. Additional in vivo studies revealed that the glycoRNA-rich membranes coated siMT1-loaded poly(lactic-co-glycolic acid) (PLGA)-polyethylene glycol (PEG) nanoparticles (GlycoRNA-NP-siMT1) effectively delivered siMT1 to AAA lesions, thereby inhibiting abdominal aortic dilation. Mechanistically, GlycoRNA-NP-siMT1 mitigated pathological remodeling of the abdominal aorta by reducing neutrophil infiltration and inhibiting the formation of NETs. Our study indicates that MT1 facilitates the progression of AAA by modulating the formation of NETs. Furthermore, GlycoRNA-NP-siMT1 show an inhibitory effect on AAA progression through a dual mechanism: they competitively inhibit neutrophil infiltration and release siMT1, which subsequently suppresses NET formation.

Polyphenols are well known for their health-related benefits, including antioxidant activities, but most of them are hydrophobic, decreasing their bioavailability. This study reports water-soluble trimeric antioxidants synthesized with l-carnosine and the hydrophobic ortho-methoxy-substituted phenolic unit, syringaldehyde or vanillin. In the DPPH assay, carnosine-syringaldehyde (7.5 μM) and carnosine-vanillin (19 μM) derivatives showed much lower IC50 values than ascorbic acid (27.5 μM) and sodium ascorbate (30.5 μM) standards. According to the AAPH assay, carnosine-syringaldehyde and carnosine-vanillin protect DNA at concentrations as low as 6.5 μM and 26 μM, respectively, while both sodium ascorbate and ascorbic acid protected until 52 μM. Another notable property of these antioxidants is that they can protect DNA well against hydroxyl radicals, produced via the Fenton reaction: carnosine-syringaldehyde showed DNA protection at all tested concentrations (833-1.6 μM), but the protection was slightly weaker between 26 and 1.6 μM. Carnosine-vanillin showed strong protection in the 833-104 μM range and some protection between 52 and 3.2 μM. Conversely, both sodium ascorbate and ascorbic acid did not protect DNA at any tested concentrations. In the pro-oxidant potential assessments, the synthesized antioxidants did not show any pro-oxidant effects at all tested concentrations. In comparison, sodium ascorbate at 833-13 μM and ascorbic acid at 833-52 μM both exhibited severe pro-oxidant effects. Our study highlights the significance of ortho-methoxy groups in antioxidants. Their electron-donating properties enhance antioxidant activities, while their steric bulk hinders the binding of transition metal ions to the phenolic hydroxyl group, thereby preventing pro-oxidant effects. The hydrophobicity of ortho-methoxy substituted phenols can be mitigated by attaching them to a highly water-soluble scaffold containing functional groups that can facilitate charge formation in the end products, such as carnosine.

This perspective discusses the essential micronutrient zinc, which functions in >3000 human proteins (the zinc proteome), and the implications of three aspects to ascertain an adequate zinc status for human health. First, the advent of highly sensitive fluorescent (bio)chemicals revealed cellular pools of zinc ions involved in signaling and secretion from cells for paracrine, autocrine, and possibly endocrine functions. Zinc signaling adds a yet unaccounted number of targeted proteins to the already impressive number of zinc proteins. Second, cellular zinc concentrations are remarkably high in the order of the concentrations of major metabolites and, therefore, at the cellular level zinc is not a trace element. Zinc is also not an antioxidant because zinc ions are redox-inactive in biology. However, zinc can express indirect pro-oxidant or proantioxidant effects depending on how cellular zinc is buffered. Zinc sites in proteins and other biomolecules can become redox-active when zinc is bound to the redox-active sulfur donor atom of cysteine. This interaction links zinc and redox metabolism, confers mobility on tightly bound zinc, and has implications for treating zinc deficiency. Third, the concept of zinc deficiency in blood as the only measure of an inadequate zinc status needs to be extended to zinc dyshomeostasis in cells because overwhelming the mechanisms controlling cellular zinc homeostasis can result in either not enough or too much available zinc. We need additional biomarkers of zinc status that determine cell-specific changes and perturbations of the system regulating cellular zinc, including functional deficits, and address the multiple genetic and environmental factors that can cause a conditioned zinc deficiency or overload. Considering the wider context of altered zinc availability in different organs, cells, and organelles impinges on whether zinc supplementation will be efficacious and adds another dimension to the already high health burden of zinc deficiency and its sequelae worldwide.

Chronic inflammatory pain is often caused by peripheral tissue damage and persistent inflammation. This disease substantially affects patients' physical and social well-being. We investigated the role of metallothionein-3 (MT3) in modulating complete Freund's adjuvant (CFA)-induced intracellular Zn2+ activity in an MT3 knockout mouse model of inflammatory pain in the hind paw. The results demonstrated that increasing intracellular Zn2+ levels ameliorate deficits in motor behavior, as well as inflammation in the paw, spleen, and thymus. Furthermore, intracellular Zn2+ was crucial in regulating oxidative stress markers (glutathione, superoxide dismutase, catalase, and malondialdehyde) and inflammatory cytokines, such as tumor necrosis factor-α and interleukin-6, in MT3 knockout mice induced with CFA. This study highlights the critical role of MT3 in coordinating the intracellular interaction with Zn2+, which is vital for the immune systems's protective functions. These interactions are fundamental for maintaining metal ion homeostasis and regulating the synthesis of various biomolecules in the body.

Intracellular metal ions play essential roles in multiple physiological processes, including catalytic action, diverse cellular processes, intracellular signaling, and electron transfer. It is crucial to maintain intracellular metal ion homeostasis which is achieved by the subtle balance of storage and release of metal ions intracellularly along with the influx and efflux of metal ions at the interface of the cell membrane. Dysregulation of intracellular metal ions has been identified as a key mechanism in triggering programmed cell death (PCD). Despite the importance of metal ions in initiating PCD, the molecular mechanisms of intracellular metal ions within these processes are infrequently discussed. An in-depth understanding and review of the role of metal ions in triggering PCD may better uncover novel tools for cancer diagnosis and therapy. Specifically, the essential roles of calcium (Ca2+), iron (Fe2+/3+), copper (Cu+/2+), and zinc (Zn2+) ions in triggering PCD are primarily explored in this review, and other ions like manganese (Mn2+/3+/4+), cobalt (Co2+/3+) and magnesium ions (Mg2+) are briefly discussed. Further, this review elaborates on the underlying chemical mechanisms and summarizes these metal ions triggering PCD in cancer therapy. This review bridges chemistry, immunology, and biology to foster the rational regulation of metal ions to induce PCD for cancer therapy.

Mammalian metallothioneins (MTs) play a crucial role in maintaining Zn(II) and Cu(I) homeostasis, as well as regulating the cellular redox potential. They are involved in cancer resistance to cisplatin-related drugs and the sequestration of toxic metal ions. To investigate their participation in specific physiological and pathological processes, it is imperative to develop an analytical method for measuring changes in protein concentration both in vitro and in vivo. Over the years, numerous methods have been developed for this purpose; however, they often suffer from issues such as low sensitivity, lengthy sample preparation, or variability in antibody batches.

In this study, we present, for the first time, the application of the fluorescent biarsenical probe CrAsH-EDT2 for the specific and ultrasensitive detection of human MTs. The interaction between the small library of biarsenical probes and 11 human MT isoforms (MT1-MT4), isolated MT2 domains, snail LlMT, and plant phytochelatin PC4 was investigated using spectrofluorimetry and mass spectrometry. A key advantage of our method is that the probe excess does not need to be removed before measurement as unbound is virtually non-fluorescent. We established protocols for MT detection in various assays, including cuvette-based, SDS-PAGE, and capillary electrophoresis. A detection limit (LOD) of 100 fg for human MT2 was achieved. Finally, we observed increased metallothionein concentration in lysates of HeLa cells treated with ZnSO4.

In conclusion, our results demonstrate that the established method is easy to apply, versatile, and can be adapted to detect metallothioneins and other poly-Cys targets in different assays and applications.

Plant metallothioneins (MTs) are low-molecular-weight proteins involved in heavy metal binding and response to stress conditions. This work aimed to analyse canola (Brassica napus L.) MTs (BnMT1-4) response to salinity and plant interaction with bacteria.

(1) We tested germination and canola growth and development in the presence of sodium chloride and bacteria Serratia plymuthica; (2) We analysed phytohormones content using LC-MS/MS; (3) We identified in silico cis-regulatory elements in promoters of BnMT1-4 genes; and (4) we investigated BnMT1-4 genes' expression in B. napus.

Under saline conditions, canola germination and plant growth were notably inhibited, whereas inoculation of seeds with S. plymuthica significantly stimulated the analysed physiological traits of B. napus. The content of auxin, abscisic acid, jasmonates, gibberellins, and salicylic acid in B. napus was significantly affected by salinity and modulated by S. plymuthica presence. The promoter regions of the BnMT1-4 genes contain numerous regulatory elements controlled by light, hormones, and various stresses. Interestingly, the expression of BnMT1-3 genes was down-regulated under salt stress, while BnMT4 transcript levels increased strongly at the highest salt concentrations with and without S. plymuthica present.

The results show that BnMT genes are differently affected by salinity and bacteria S. plymuthica and significantly correlate with particular phytohormones content in canola tissues, confirming the diversified functions of MTs in plant responses to changing environment.

Zinc is an essential micronutrient with a myriad of key roles in human health. This review summarizes mechanistic data supporting the protective effects of zinc on metal toxicity and discusses the framework for an interventional clinical trial of zinc supplementation within a metal exposed Native American community.

Many metals have common underlying mechanisms of toxicity that contribute to adverse human health effects. Studies demonstrate that multiple aspects of metal toxicity can be attributed to disruption of essential zinc-dependent functions. Multiple lines of evidence suggest that zinc may confer protection against metal toxicity in human populations with mixed-metal exposures. Thinking Zinc is a mechanism-informed intervention study of zinc supplementation to test the potential benefits of zinc while maintaining a culturally responsive research approach. The current knowledge of diverse metal and zinc interactions, coupled with strong mechanistic evidence for zinc benefits in the context of toxic metal exposures, supports the hypothesis that zinc supplementation may mitigate the impact of toxic metals exposures in populations with chronic mixed metal exposures and in populations with low zinc status.

In the past decade, evidence for the numerous roles of copper (Cu) in mammalian physiology has grown exponentially. The discoveries of Cu involvement in cell signaling, autophagy, cell motility, differentiation, and regulated cell death (cuproptosis) have markedly extended the list of already known functions of Cu, such as a cofactor of essential metabolic enzymes, a protein structural component, and a regulator of protein trafficking. Novel and unexpected functions of Cu transporting proteins and enzymes have been identified, and new disorders of Cu homeostasis have been described. Significant progress has been made in the mechanistic studies of two classic disorders of Cu metabolism, Menkes disease and Wilson's disease, which paved the way for novel approaches to their treatment. The discovery of cuproptosis and the role of Cu in cell metastatic growth have markedly increased interest in targeting Cu homeostatic pathways to treat cancer. In this review, we summarize the established concepts in the field of mammalian Cu physiology and discuss how new discoveries of the past decade expand and modify these concepts. The roles of Cu in brain metabolism and in cell functional speciation and a recently discovered regulated cell death have attracted significant attention and are highlighted in this review.

Proteins are dynamic entities that adopt diverse conformations, which play a pivotal role in their function. Understanding these conformations is essential, and protein collective motions, particularly those captured by normal mode (NM) and their linear combinations, provide a robust means for conformational sampling. This work introduces a novel approach to obtaining a uniformly oriented set of a given number of lowest frequency NM combined vectors and generating harmonically equidistant restrained structures along them. They are all thus uniformly located on concentric hyperspheres, systematically covering the defined NM space fully. The generated structures are further relaxed with standard molecular dynamics (MD) simulations to explore the conformational space. The efficiency of the approach we termed "distributed points Molecular Dynamics using Normal Modes" (dpMDNM) was assessed by applying it to hen egg-white lysozyme and human cytochrome P450 3A4 (CYP3A4). To this purpose, we compared our new approach with other methods and analyzed the sampling of existing experimental structures. Our results demonstrate the efficacy of dpMDNM in extensive conformational sampling, particularly as more NMs are considered. Ensembles generated by dpMDNM exhibited a broad coverage of the experimental structures, providing valuable insights into the functional aspects of lysozyme and CYP3A4. Furthermore, dpMDNM also covered lysozyme structures with relatively elevated energies corresponding to transient states not easily obtained by standard MD simulations, in conformity with nuclear magnetic resonance structural indications. This method offers an efficient and rational framework for comprehensive protein conformational sampling, contributing significantly to our understanding of protein dynamics and function.

Zinc (Zn) is an essential but toxic trace element and is widely available in the natural environment. In the present study, we developed a re-absorption physiologically based pharmacokinetic (PBPK) model based on long-term dietary exposure to gain insights into the physiological mechanisms of uptake, tissue distribution, storage, and excretion of Zn in marine juvenile gilt-head breams Sparus aurata (with stomach). The PBPK model incorporated the kinetic processes of Zn transfer from fish liver to gastrointestinal system and used the Markov Monte Carlo algorithm to estimate the distribution of model parameters. The model fit indicated that the stomach and intestine of fish were key organs in regulating the concentration of Zn entering the internal environment, with excess exogenous Zn (120 mg/kg) being excreted in feces (rate constant of 5.23 d-1). Modeling results also indicated that liver (3.00 d-1), spleen (1.41 d-1) and kidney (0.51 d-1) were the main tissues responding to blood Zn flux by accumulation and detoxification. Fish kidneys exposed to 60 mg/kg and 120 mg/kg Zn had different regenerative capacities, resulting in different detoxification functions. A higher dietary Zn (120 mg/kg) disrupted the intestinal reabsorption process in marine fish. This study showed that exogenous Zn was directly accumulated in organs through the gastrointestinal-hepatic system, which is an important pathways for regulating metal homeostasis in marine fish. The results provided important understanding of the mechanisms of metal regulation and transport in marine fish.

The essential trace element, zinc, regulates virtually all aspects of cellular physiology, particularly cell proliferation and survival. Diverse families of metal transporters, metallothioneins, and metal-responsive transcriptional regulators are linked to zinc homeostasis. However, the mechanism underlying the regulation of systemic zinc homeostasis remains largely unknown. Here, it is reported that the intestinal transporter SLC30A1 plays an essential role in maintaining systemic zinc homeostasis. Using several lines of tissue-specific knockout mice, it is found that intestinal Slc30a1 plays a critical role in survival. Furthermore, lineage tracing reveals that Slc30a1 is localized to the basolateral membrane of intestinal epithelial cells (IECs). It is also found that Slc30a1 safeguards both intestinal barrier integrity and systemic zinc homeostasis. Finally, an integrative analysis of the cryo-EM structure and site-specific mutagenesis of human SLC30A1 are performed and a zinc transport mechanism of SLC30A1 unique within the SLC30A family, with His43 serving as a critical residue for zinc selectivity, is identified.

Cancer has a devastating impact globally regardless of gender, age, and community, which continues its severity to the population due to the lack of efficient strategy for the cancer diagnosis and treatment. According to the World Health Organisation report, one out of six people dies due to this deadly cancer and we need effective strategies to regulate it. In this context, trace element has a very hidden and unexplored role and require more attention from investigators. The variation in concentration of trace elements was observed during comparative studies on a cancer patient and a healthy person making them an effective target for cancer regulation. The percentage of trace elements present in the human body depends on environmental exposure, food habits, and habitats and could be instrumental in the early diagnosis of cancer. In this review, we have conducted inclusive analytics on trace elements associated with the various types of cancers and explored the several methods involved in their analysis. Further, intricacies in the correlation of trace elements with prominent cancers like prostate cancer, breast cancer, and leukaemia are represented in this review. This comprehensive information on trace elements proposes their role during cancer and as biomarkers in cancer diagnosis.

In this study, we performed synchrotron-based micro-X-ray fluorescence (μ-XRF) imaging of elements Zn and S, and X-ray absorption near edge spectroscopy (XANES) coupled with μ-XRF for identification of Zn and S species in the condylar zones of a rat temporomandibular joint (TMJ). Histologic localization of Zn and hypoxia-inducible factor-1α (HIF-1α) were mapped using an optical microscope. These data were visually correlated with μ-XRF and XANES data to provide insights into plausible biological S-species in Z-enriched condylar zones of a rat TMJ. Furthermore, μ-XRF coupled with micro-X-ray diffraction (μ-XRD) was used to underline Z-incorporated biological apatite in the subchondral bone and bone of the rat TMJ. Results illustrated the potential dependence between biometal Zn and nonmetal S and their collective governance of cell and tissue functions in a zone-specific manner. Elemental Zn with organic and inorganic S-species at the cartilage-bone interface and transformation of plausible Zn-enriched mineralization kinetics of biological apatite from subchondral bone to condylar bone were ascertained using μ-XRF-XANES and μ-XRD. The coupled μ-XRF-XANES complementing with μ-XRD and immunohistology provided an informative view of S and Zn and their association with zone-specific biological pathways in situ. Understanding the spatial distributions of the main S-species with redox-inert Zn in regions of cartilage, bone, and the interface is essential for further unlocking questions surrounding formation and resorption-related biomineralization pathways as related to osteoarthritis or genetically inherited diseases. Using these complementary techniques with microspectroscopic spatial information provided insights into the associations between biometal Zn and nonmetal S and a window into detecting the plausible early-stage diagnostic biomarkers for humans with TMJ osteoarthritis.

Metallothioneins (MTs) are small proteins that coordinate d-block metal ions in sulfur-metal clusters to control metal ion concentrations within the cell. Here we study metal cluster formation in the MT of the periwinkle Littorina littorea (LlMT) by nuclear magnetic resonance (NMR). We demonstrate that the three Cd2+ ions in each domain are taken up highly cooperatively, that is, in an all-or-none fashion, with a four- to six-fold higher affinity for the C-terminal domain. During the transfer of metal ions from Cd2+-loaded MT to apo MT, Cd2+ is most efficiently transferred from the metalated protein to the apo C-terminal domain. This behavior might be connected to unique structural motifs in the C-terminal domain, such as two double-CXC motifs and an increased proportion of positively charged residues. In Cd2+/Zn2+ metal exchange experiments, the N-terminal domain displayed the most efficient inter-molecular metal exchange. Amide hydrogen exchange reveals fewer protected amides for the N-terminal domain, suggesting the structure might more easily "open up" to facilitate metal exchange. Experiments with a physical separation of donor and acceptor species demonstrate that metal exchange and transfer require protein-protein contacts. These findings provide insights into the mechanism of metal uptake and metal transfer, which are important processes during metal detoxification in snail MTs.

The study investigated the performance of a novel magnetic hybrid MIL-53(Fe)/Fe3O4@TiO2 composite for removing reactive red 195 (RR195) dye from water using UVc light. Various analytical techniques were used to characterize the nanocomposite materials. X-ray diffraction analysis confirmed the presence of MIL-53(Fe) and TiO2 in the composite. FT-IR analysis identified carboxyl and Ti-O-Ti groups in the photocatalyst structure. The study evaluated the effects of pH, dye concentration, photocatalyst dosage, and temperature on RR195 photodegradation. The Langmuir-Hinshelwood kinetic model provided the best fit for the reaction rate. Optimal conditions for an 84 % dye degradation were found at a photocatalyst dose of 15 mg/100 mL, pH 3, dye concentration of 100 mg/L, and 35 °C after 120 minutes of UVc light exposure. Thermodynamic analysis indicated an endothermic reaction with positive values for Δ#H and negative values for Δ#S. The MIL-53(Fe)/Fe3O4@TiO2 composite demonstrated excellent stability and achieved over 90 % dye degradation after five cycles. Overall, the composite shows promise for treating wastewater with dyes.

The small Cys-rich protein metallothionein (MT) binds several metal ions in clusters within two domains. While the affinity of MT for both toxic and essential metals has been well studied, the thermodynamics of this binding has not. We have used isothermal titration calorimetry measurements to quantify the change in enthalpy (ΔH) and change in entropy (ΔS) when metal ions bind to the two ubiquitous isoforms of MT. The seven Zn2+ that bind sequentially at pH 7.4 do so in two populations with different coordination thermodynamics, an initial four that bind randomly with individual tetra-thiolate coordination and a subsequent three that bind with bridging thiolate coordination to assemble the metal clusters. The high affinity of MT for both populations is due to a very favourable binding entropy that far outweighs an unfavourable binding enthalpy. This originates from a net enthalpic penalty for Zn2+ displacement of protons from the Cys thiols and a favourable entropic contribution from the displaced protons. The thermodynamics of other metal ions binding to MT were determined by their displacement of Zn2+ from Zn7MT and subtraction of the Zn2+-binding thermodynamics. Toxic Cd2+, Pb2+, and Ag+, and essential Cu+, also bind to MT with a very favourable binding entropy but a net binding enthalpy that becomes increasingly favourable as the metal ion becomes a softer Lewis acid. These thermodynamics are the origin of the high affinity, selectivity, and domain specificity of MT for these metal ions and the molecular basis for their in vivo binding competition.

The mutual relationship between peptides and metal ions enables metalloproteins to have crucial roles in biological systems, including structural, sensing, electron transport, and catalytic functions. The effort to reproduce or/and enhance these roles, or even to create unprecedented functions, is the focus of protein design, the first step toward the comprehension of the complex machinery of nature. Nowadays, protein design allows the building of sophisticated scaffolds, with novel functions and exceptional stability. Recent progress in metalloprotein design has led to the building of peptides/proteins capable of orchestrating the desired functions of different metal cofactors. The structural diversity of peptides allows proper selection of first- and second-shell ligands, as well as long-range electrostatic and hydrophobic interactions, which represent precious tools for tuning metal properties. The scope of this review is to discuss the construction of metal sites in de novo designed and miniaturized scaffolds. Selected examples of mono-, di-, and multi-nuclear binding sites, from the last 20 years will be described in an effort to highlight key artificial models of catalytic or electron-transfer metalloproteins. The authors' goal is to make readers feel like guests at the marriage between peptides and metal ions while offering sources of inspiration for future architects of innovative, artificial metalloproteins.

Sinopotamon Henanense expresses two metal‒induced metallothioneins (MTs), Cd‒induced MT and Cu‒induced MT (ShCuMT). The Cd‒induced MT has been characterized as a Cd‒thiolate MT. However, it is unknown whether ShCuMT is a Cu‒thiolate MT. In the present study, ShCuMT was expressed heterologously in Escherichia coli and purified by Ni‒NTA column and superdex‒75 column. And its metal‒binding feature was evaluated by DTNB reaction, circular dichroism spectroscopy (CD), isothermal microtitration (ITC), electrospray flight mass spectrometry (ESI‒TOF‒MS), and matrix‒assisted laser desorption ionization flight mass spectrometry (MALDI‒TOF‒MS). Bioinformatics analysis demonstrated that ShCuMT possessed the cysteine‒triplet motif of a Cu‒specific MT. Expression and purification of ShCuMT illustrated that SUMO tag used as the production system for ShCuMT resulted in a high production yield. The stability order of ShCuMT binding metal ions were Cu (Ⅰ) > Cd (Ⅱ) > Zn (Ⅱ). The CD spectrum indicated that ShCuMT binding with Cu (I) exhibited a compact thiol metal clusters structure. Besides, there emerged no a visible nickel‒thiol absorption after Ni‒NTA column affinity chromatography. The ITC results implied that Cu‒ShCuMT possessed the optimal thermodynamic conformation and the highest stoichiometric number of Cu (Ⅰ). Overall, the results suggested that SUMO fusion system is a robust and inexpensive approach for ShCuMT expression and Ni‒NTA column had no influence on metal binding of ShCuMT and Cu(Ⅰ) was considered its cognate metal ion, and ShCuMT possessed canonical Cu‒thiolate characteristics. The metal binding feature of ShCuMT reported here contributes to elucidating the structure‒function relationship of ShCuMT in S. Henanense.

Increased awareness of the impact of human activities on the environment has emerged in recent decades. One significant global environmental and human health issue is the development of materials that could potentially have negative effects. These materials can accumulate in the environment, infiltrate organisms, and move up the food chain, causing toxic effects at various levels. Therefore, it is crucial to assess materials comprising nano-scale particles due to the rapid expansion of nanotechnology. The aquatic environment, particularly vulnerable to waste pollution, demands attention. This review provides an overview of the behavior and fate of metallic nanoparticles (NPs) in the aquatic environment. It focuses on recent studies investigating the toxicity of different metallic NPs on aquatic organisms, with a specific emphasis on thiol-biomarkers of oxidative stress such as glutathione, thiol- and related-enzymes, and metallothionein. Additionally, the selection of suitable measurement methods for monitoring thiol-biomarkers in NPs' ecotoxicity assessments is discussed. The review also describes the analytical techniques employed for determining levels of oxidative stress biomarkers.

"Double scale" is a poorly characterized skin defect of crocodilians that drastically reduces the economic value of crocodilian skin. This study investigated the morphology and pathogenesis of double scale in a ranching farm of American alligators (Alligator mississippiensis). We compared the histopathology of skin and selected organs (liver, lung, kidney, heart, spleen, intestine, and brain) of alligators with double scale against healthy control animals, together with serum and liver vitamin and mineral levels. Skin affected with double scale had statistically significant hyperkeratosis, epidermal atrophy, and increased basal cell degeneration compared with control alligators (P < .0001). Interestingly, all alligators with double scale had varying degrees of hepatic fibrosis. Feed analysis showed that alligators that had double scale and hepatic fibrosis had prolonged dietary exposure to high levels of vitamin A, iron, and copper. Serum analysis indicated that levels of zinc (p < .0001), copper (P < .05), and vitamin E (P < .002) were significantly lower in alligators with hepatic fibrosis and double scale compared with controls. Finally, immunohistochemical analysis of skin with double scale showed a marked reduction in immunolabeling with the zinc-binding protein metallothionein. These results suggest that zinc deficiency, in combination with other micronutrient anomalies, may play a role in the pathogenesis of double scale in alligators with liver fibrosis.

Within the intricate landscape of the proteome, approximately 30% of all proteins bind metal ions. This repertoire is even larger when considering all the different forms of a protein, known as proteoforms. Here, we propose the term "metalloforms" to refer to different structural or functional variations of a protein resulting from the binding of various hetero- or homogeneous metal ions. Using human Cu(I)/Zn(II)-metallothionein-3 as a representative model, we developed a chemical proteomics strategy to simultaneously differentiate and map Zn(II) and Cu(I) metal binding sites. In the first labeling step, N-ethylmaleimide reacts with Cysteine (Cys), resulting in the dissociation of all Zn(II) ions while Cu(I) remains bound to the protein. In the second labeling step, iodoacetamide is utilized to label Cu(I)-bound Cys residues. Native mass spectrometry (MS) was used to determine the metal/labeling protein stoichiometries, while bottom-up/top-down MS was used to map the Cys-labeled residues. Next, we used a developed methodology to interrogate an isolated rabbit liver metallothionein fraction containing three metallothionein-2 isoforms and multiple Cd(II)/Zn(II) metalloforms. The approach detailed in this study thus holds the potential to decode the metalloproteoform diversity within other proteins.

Bone homeostasis is maintained by an intricate balance between osteoclasts and osteoblasts, which becomes disturbed in osteoporosis. Metallothioneins (MTs) are major contributors in cellular zinc regulation. However, the role of MTs in bone cell regulation has remained unexplored. Single-cell RNA sequencing analysis discovered that, unlike the expression of other MT members, the expression of MT3 was unique to osteoclasts among various macrophage populations and was highly upregulated during osteoclast differentiation. This unique MT3 upregulation was validated experimentally and supported by ATAC sequencing data analyses. Downregulation of MT3 by gene knockdown or knockout resulted in excessive osteoclastogenesis and exacerbated bone loss in ovariectomy-induced osteoporosis. Transcriptome sequencing of MT3 knockdown osteoclasts and gene set enrichment analysis indicated that the oxidative stress and redox pathways were enriched, which was verified by MT3-dependent regulation of reactive oxygen species (ROS). In addition, MT3 deficiency increased the transcriptional activity of SP1 in a manner dependent on intracellular zinc levels. This MT3-zinc-SP1 axis was crucial for the control of osteoclasts, as zinc chelation and SP1 knockdown abrogated the promotion of SP1 activity and osteoclastogenesis by MT3 deletion. Moreover, SP1 bound to the NFATc1 promoter, and overexpression of an inactive SP1 mutant negated the effects of MT3 deletion on NFATc1 and osteoclastogenesis. In conclusion, MT3 plays a pivotal role in controlling osteoclastogenesis and bone metabolism via dual axes involving ROS and SP1. The present study demonstrated that MT3 elevation is a potential therapeutic strategy for osteolytic bone disorders, and it established for the first time that MT3 is a crucial bone mass regulator.

Cuproptosis, a newly characterized form of regulated cell death driven by copper accumulation, has emerged as a significant mechanism underlying various non-cancerous diseases. This review delves into the complex interplay between copper metabolism and the pathogenesis of conditions such as Wilson's disease (WD), neurodegenerative disorders, and cardiovascular pathologies. We examine the molecular mechanisms by which copper dysregulation induces cuproptosis, highlighting the pivotal roles of key copper transporters and enzymes. Additionally, we evaluate the therapeutic potential of copper chelation strategies, which have shown promise in experimental models by mitigating copper-induced cellular damage and restoring physiological homeostasis. Through a comprehensive synthesis of recent advancements and current knowledge, this review underscores the necessity of further research to translate these findings into clinical applications. The ultimate goal is to harness the therapeutic potential of targeting cuproptosis, thereby improving disease management and patient outcomes in non-cancerous conditions associated with copper dysregulation.

Prostate, bladder, and kidney cancers are the most common malignancies of the urinary system. Chemotherapeutic drugs are generally used as adjuvant treatment in the middle, late, or recurrence stages after surgery for urologic cancers. However, traditional chemotherapy is plagued by problems such as poor efficacy, severe side effects, and complications. Copper-containing nanomedicines are promising novel cancer treatment modalities that can potentially overcome these disadvantages. Copper homeostasis and cuproptosis play crucial roles in the development, adaptability, and therapeutic sensitivity of urological malignancies. Cuproptosis refers to the direct binding of copper ions to lipoylated components of the tricarboxylic acid cycle, leading to protein oligomerization, loss of iron-sulfur proteins, proteotoxic stress, and cell death. This review focuses on copper homeostasis and cuproptosis as well as recent findings on copper and cuproptosis in urological malignancies. Furthermore, we highlight the potential therapeutic applications of copper- and cuproptosis-targeted therapies to better understand cuproptosis-based drugs for the treatment of urological tumors in the future.

Herby, the interaction of metallothioneins with commonly used Pt-based anticancer drugs - cisplatin, carboplatin, and oxaliplatin - was investigated using the combined power of elemental (i.e. LA-ICP-MS, CE-ICP-MS) and molecular (i.e. MALDI-TOF-MS) analytical techniques providing not only required information about the interaction, but also the benefit of low sample consumption. The amount of Cd and Pt incorporated within the protein was determined for protein monomers and dimer/oligomers formed by non-oxidative dimerization. Moreover, fluorescence spectrometry using Zn2+-selective fluorescent indicator - FluoZin3 - was employed to monitor the ability of Pt drugs to release natively occurring Zn from the protein molecule. The investigation was carried out using two protein isoforms (i.e. MT2, MT3), and significant differences in behaviour of these two isoforms were observed. The main attention was paid to elucidating whether the protein dimerization/oligomerization may be the reason for the potential failure of the anticancer therapy based on these drugs. Based on the results, it was demonstrated that the interaction of MT2 (both monomers and dimers) interacted with Pt drugs significantly less compared to MT3 (both monomers and dimers). Also, a significant difference between monomeric and dimeric forms (both MT2 and MT3) was not observed. This may suggest that dimer formation is not the key factor leading to the inactivation of Pt drugs.

Copper is a critical trace element in biological systems due the vast number of essential enzymes that require the metal as a cofactor, including cytochrome c oxidase, superoxide dismutase and dopamine-β-hydroxylase. Due its key role in oxidative metabolism, antioxidant defence and neurotransmitter synthesis, copper is particularly important for neuronal development and proper neuronal function. Moreover, increasing evidence suggests that copper also serves important functions in synaptic and network activity, the regulation of circadian rhythms, and arousal. However, it is important to note that because of copper's ability to redox cycle and generate reactive species, cellular levels of the metal must be tightly regulated to meet cellular needs while avoiding copper-induced oxidative stress. Therefore, it is essential that the intricate system of copper transporters, exporters, copper chaperones and copper trafficking proteins function properly and in coordinate fashion. Indeed, disorders of copper metabolism such as Menkes disease and Wilson disease, as well as diseases linked to dysfunction of copper-requiring enzymes, such as SOD1-linked amyotrophic lateral sclerosis, demonstrate the dramatic neurological consequences of altered copper homeostasis. In this review, we explore the physiological importance of copper in the nervous system as well as pathologies related to improper copper handling.

Highly colored azo dye-contaminated wastewater poses significant environmental threats and requires effective treatment before discharge. The anaerobic azo dye treatment method is a cost-effective and environmentally friendly solution, while its time-consuming and inefficient processes present substantial challenges for industrial scaling. Thus, the use of iron materials presents a promising alternative. Laboratory studies have demonstrated that systems coupled with iron materials enhance the decolorization efficiency and reduce the processing time. To fully realize the potential of iron materials for anaerobic azo dye treatment, a comprehensive synthesis and evaluation based on individual-related research studies, which have not been conducted to date, are necessary. This review provides, for the first time, an extensive and detailed overview of the utilization of iron materials for azo dye treatment, with a focus on decolorization. It assesses the treatment potential, analyzes the influencing factors and their impacts, and proposes metabolic pathways to enhance anaerobic dye treatment using iron materials. The physicochemical characteristics of iron materials are also discussed to elucidate the mechanisms behind the enhanced bioreduction of azo dyes. This study further addresses the current obstacles and outlines future prospects for industrial-scale application of iron-coupled treatment systems.

The past few decades have witnessed a rapid growth in the prevalence of nonalcoholic fatty liver disease (NAFLD). While the ketogenic diet (KD) is considered for managing NAFLD, the safety and efficacy of the KD on NAFLD has been a controversial topic. Here, we aimed to investigate the effect of KD of different durations on metabolic endpoints in mice with NAFLD and explore the underlying mechanisms.

NAFLD mice were fed with KD for 1, 2, 4 and 6 weeks, respectively. The blood biochemical indexes (blood lipids, AST, ALT and etc.) and liver fat were measured. The LC-MS/MS based proteomic analysis was performed on liver tissues. Metallothionein-2 (MT2) was knocked down with adeno-associated virus (AAV) or small interfering RNA (siRNA) in NAFLD mice and AML-12 cells, respectively. H&E, BODIPY and ROS staining were performed to examine lipid deposition and oxidative stress. Furthermore, MT2 protein levels, nucleus/cytoplasm distribution and DNA binding activity of peroxisome proliferators-activated receptors α (PPARα) were evaluated.

KD feeding for 2 weeks showed the best improvement on NAFLD phenotype. Proteomic analysis revealed that MT2 was a key candidate for different metabolic endpoints of NAFLD affected by different durations of KD feeding. MT2 knockdown in NAFLD mice blocked the effects of 2 weeks of KD feeding on HFD-induced steatosis. In mouse primary hepatocytes and AML-12 cells, MT2 protein levels were induced by β-hydroxybutyric acid (β-OHB). MT2 Knockdown blunted the effects of β-OHB on alleviating PA-induced lipid deposition. Mechanistically, 2 weeks of KD or β-OHB treatment reduced oxidative stress and upregulated the protein levels of MT2 in nucleus, which subsequently increased its DNA binding activity and PPARα protein expression.

Collectively, these findings indicated that KD feeding prevented NAFLD in a time dependent manner and MT2 is a potential target contributing to KD improvement on steatosis.

Oligodendrocytes develop through sequential stages and understanding pathways regulating their differentiation remains an important area of investigation. Zinc is required for the function of enzymes, proteins and transcription factors, including those important in myelination and mitosis. Our previous studies using the ratiometric zinc sensor chromis-1 demonstrated a reduction in intracellular free zinc concentrations in mature MBP+ oligodendrocytes compared with earlier stages (Bourassa et al., 2018). We performed a more detailed developmental study to better understand the temporal course of zinc homeostasis across the oligodendrocyte lineage. Using chromis-1, we found a transient increase in free zinc after O4+,O1- pre-oligodendrocytes were switched from proliferation medium into terminal differentiation medium. To gather other evidence for dynamic regulation of free zinc during oligodendrocyte development, qPCR was used to evaluate mRNA expression of major zinc storage proteins metallothioneins (MTs) and metal regulatory transcription factor 1 (MTF1), which controls expression of MTs. MT1, MT2 and MTF1 mRNAs were increased several fold in mature oligodendrocytes compared to oligodendrocytes in proliferation medium. To assess the depth of the zinc buffer, we assayed zinc release from intracellular stores using the oxidizing thiol reagent 2,2'-dithiodipyridine (DTDP). Exposure to DTDP resulted in ∼ 100% increase in free zinc in pre-oligodendrocytes but, paradoxically more modest ∼ 60% increase in mature oligodendrocytes despite increased expression of MTs. These results suggest that zinc homeostasis is regulated during oligodendrocyte development, that oligodendrocytes are a useful model for studying zinc homeostasis in the central nervous system, and that regulation of zinc homeostasis may be important in oligodendrocyte differentiation.

In this pilot study on subway workers, we explored the relationships between particle exposure and oxidative stress biomarkers in exhaled breath condensate (EBC) and urine to identify the most relevant biomarkers for a large-scale study in this field.

We constructed a comprehensive occupational exposure assessment among subway workers in three distinct jobs over 10 working days, measuring daily concentrations of particulate matter (PM), their metal content and oxidative potential (OP). Individual pre- and post-shift EBC and urine samples were collected daily. Three oxidative stress biomarkers were measured in these matrices: malondialdehyde (MDA), 8-hydroxy-2'deoxyguanosine (8-OHdG) and 8-isoprostane. The association between each effect biomarker and exposure variables was estimated by multivariable multilevel mixed-effect models with and without lag times.

The OP was positively associated with Fe and Mn, but not associated with any effect biomarkers. Concentration changes of effect biomarkers in EBC and urine were associated with transition metals in PM (Cu and Zn) and furthermore with specific metals in EBC (Ba, Co, Cr and Mn) and in urine (Ba, Cu, Co, Mo, Ni, Ti and Zn). The direction of these associations was both metal- and time-dependent. Associations between Cu or Zn and MDAEBC generally reached statistical significance after a delayed time of 12 or 24 h after exposure. Changes in metal concentrations in EBC and urine were associated with MDA and 8-OHdG concentrations the same day.

Associations between MDA in both EBC and urine gave opposite response for subway particles containing Zn versus Cu. This diverting Zn and Cu pattern was also observed for 8-OHdG and urinary concentrations of these two metals. Overall, MDA and 8-OHdG responses were sensitive for same-day metal exposures in both matrices. We recommend MDA and 8-OHdG in large field studies to account for oxidative stress originating from metals in inhaled particulate matter.

The relationship between dietary metal intake and mortality risk is controversial, and we investigated the relationship between intake of five metals (iron, copper, selenium, zinc, and magnesium) and all-cause, cardiovascular mortality in the total population, gender subgroups, and age subgroups.

17,207 participants from the National Health and Nutrition Examination Survey (NHANES) database from 2009 to 2016 were included in this study. Kaplan-Meier survival curves, multivariate Cox proportional hazards models, and restrictive cubic spline (RCS) curves were used to explore the association between metal intake and all-cause, cardiovascular mortality.

In this study, the average dietary metal intake of men and older people was lower than that of women and younger people. The RCS curves found in the whole population that all-cause mortality was negative linearly associated with copper intakes, L-shaped with zinc and magnesium intakes. Further subgroup analyses of copper, zinc, and magnesium by age and gender revealed that only magnesium showed statistically significant differences in the age subgroups. In the 20-40 population, there was a non-linear increasing trend in magnesium intake and all-cause mortality, whereas there was a non-linear decreasing trend in the > 60 population.

The relationship between metal intake and mortality is more than a simple linear correlation, and differences in age can affect this correlation. In metal exposure studies, different populations can be studied to better determine the effect of metal exposure on mortality.

The dataset used for statistical analysis in this study is available on the NHANES website: https://www.cdc.gov/nchs/nhanes/index.htm.

Low-complexity domains (LCDs) in proteins are typically enriched in one or two predominant amino acids. As a result, LCDs often exhibit unusual structural/biophysical tendencies and can occupy functional niches. However, for each organism, protein sequences must be compatible with intracellular biomolecules and physicochemical environment, both of which vary from organism to organism. This raises the possibility that LCDs may occupy sequence spaces in select organisms that are otherwise prohibited in most organisms. Here, we report a comprehensive survey and functional analysis of LCDs in all known reference proteomes (>21k organisms), with added focus on rare and unusual types of LCDs. LCDs were classified according to both the primary amino acid and secondary amino acid in each LCD sequence, facilitating detailed comparisons of LCD class frequencies across organisms. Examination of LCD classes at different depths (i.e., domain of life, organism, protein, and per-residue levels) reveals unique facets of LCD frequencies and functions. To our surprise, all 400 LCD classes occur in nature, although some are exceptionally rare. A number of rare classes can be defined for each domain of life, with many LCD classes appearing to be eukaryote-specific. Certain LCD classes were consistently associated with identical functions across many organisms, particularly in eukaryotes. Our analysis methods enable simultaneous, direct comparison of all LCD classes between individual organisms, resulting in a proteome-scale view of differences in LCD frequencies and functions. Together, these results highlight the remarkable diversity and functional specificity of LCDs across all known life forms.

The objective of the current review is to summarize the current state of optical methods in redox biology. It consists of two parts, the first is dedicated to genetically encoded fluorescent indicators and the second to Raman spectroscopy. In the first part, we provide a detailed classification of the currently available redox biosensors based on their target analytes. We thoroughly discuss the main architecture types of these proteins, the underlying engineering strategies for their development, the biochemical properties of existing tools and their advantages and disadvantages from a practical point of view. Particular attention is paid to fluorescence lifetime imaging microscopy as a possible readout technique, since it is less prone to certain artifacts than traditional intensiometric measurements. In the second part, the characteristic Raman peaks of the most important redox intermediates are listed, and examples of how this knowledge can be implemented in biological studies are given. This part covers such fields as estimation of the redox states and concentrations of Fe-S clusters, cytochromes, other heme-containing proteins, oxidative derivatives of thiols, lipids, and nucleotides. Finally, we touch on the issue of multiparameter imaging, in which biosensors are combined with other visualization methods for simultaneous assessment of several cellular parameters.

Cellular and organismic copper (Cu) homeostasis is regulated by Cu transporters and Cu chaperones to ensure the controlled uptake, distribution and export of Cu ions. Many of these processes have been extensively investigated in mammalian cell culture, as well as in humans and in mammalian model organisms. Most of the human genes encoding proteins involved in Cu homeostasis have orthologs in the model organism, Caenorhabditis elegans (C. elegans). Starting with a compilation of human Cu proteins and their orthologs, this review presents an overview of Cu homeostasis in C. elegans, comparing it to the human system, thereby establishing the basis for an assessment of the suitability of C. elegans as a model to answer mechanistic questions relating to human Cu homeostasis.

Understanding the homeostatic interactions among essential trace metals is important for explaining their roles in cellular systems. Recent studies in vertebrates suggest that cellular Mn metabolism is related to Zn metabolism in multifarious cellular processes. However, the underlying mechanism remains unclear. In this study, we examined the changes in the expression of proteins involved in cellular Zn and/or Mn homeostatic control and measured the Mn as well as Zn contents and Zn enzyme activities to elucidate the effects of Mn and Zn homeostasis on each other. Mn treatment decreased the expression of the Zn homeostatic proteins metallothionein (MT) and ZNT1 and reduced Zn enzyme activities, which were attributed to the decreased Zn content. Moreover, loss of Mn efflux transport protein decreased MT and ZNT1 expression and Zn enzyme activity without changing extracellular Mn content. This reduction was not observed when supplementing with the same Cu concentrations and in cells lacking Cu efflux proteins. Furthermore, cellular Zn homeostasis was oppositely regulated in cells expressing Zn and Mn importer ZIP8, depending on whether Zn or Mn concentration was elevated in the extracellular milieu. Our results provide novel insights into the intricate interactions between Mn and Zn homeostasis in mammalian cells and facilitate our understanding of the physiopathology of Mn, which may lead to the development of treatment strategies for Mn-related diseases in the future.

Metallothioneins (MTs) have a strong affinity for zinc (Zn) and remain at a sufficiently high level in mitochondria. As the avian embryo is highly susceptible to oxidative damage and relatively easy to manipulate in a naturally closed chamber, it is an ideal model of the effects of oxidative stress on mitochondrial function. However, the protective roles and molecular mechanisms of Zn-inducible protein expression on mitochondrial function in response to various stressors are poorly understood.

The study aimed to investigate the mechanisms by which Zn-induced MT4 expression protects mitochondrial function and energy metabolism subjected to oxidative stress using the avian embryo and embryonic primary hepatocyte models.

First, we investigated whether MT4 expression alters mitochondrial function. Then, we examined the effects of Zn-induced MT4 overexpression and MT4 silencing on embryonic primary hepatocytes from breeder hens fed a normal Zn diet subjected to a tert-butyl hydroperoxide (BHP) oxidative stress challenge during incubation. In vivo, the avian embryos from hens fed the Zn-deficient and Zn-adequate diets were used to determine the protective roles of Zn-induced MT4 expression on the function of mitochondria exposed to oxidative stress induced by in ovo BHP injection.

An in vitro study revealed that Zn-induced MT4 expression reduced reactive oxygen species accumulation in primary hepatocytes. MT4 silencing exacerbated BHP-mediated mitochondrial dysfunction whereas Zn-inducible MT4 overexpression mitigated it. Another in vivo study disclosed that maternal Zn-induced MT4 expression protected mitochondrial function in chick embryo hepatocytes against oxidative stress by inhibiting the peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α)/peroxisome proliferators-activated receptor-γ (PPAR-γ) pathway.

This study underscores the potential protective roles of Zn-induced MT4 expression via the downregulation of the PGC-1α/PPAR-γ pathway on mitochondrial function stimulated by the stress challenge in the primary hepatocytes in an avian embryo model. Our findings suggested that Zn-induced MT4 expression could provide a new therapeutic target and preventive strategy for repairing mitochondrial dysfunction in disease.

The chemical properties of toxic cadmium and essential zinc are very similar, and organisms require intricate mechanisms that drive selective handling of metals. Previously regarded as unspecific "metal sponges", metallothioneins (MTLs) are emerging as metal selectivity filters. By utilizing C. elegans mtl-1 and mtl-2 knockout strains, metal accumulation in single worms, single copy fluorescent-tagged transgenes, isoform specific qPCR and lifespan studies it was possible to demonstrate that the handling of cadmium and zinc by the two C. elegans metallothioneins differs fundamentally: the MTL-2 protein can handle both zinc and cadmium, but when it becomes unavailable, either via a knockout or by elevated cadmium exposure, MTL-1 takes over zinc handling, leaving MTL-2 to sequester cadmium. This division of labour is reflected in the folding behaviour of the proteins: MTL-1 folded well in presence of zinc but not cadmium, the reverse was the case for MTL-2. These differences are in part mediated by a zinc-specific mononuclear His3Cys site in the C-terminal insertion of MTL-1; its removal affected the entire C-terminal domain and may shift its metal selectivity towards zinc. Overall, we uncover how metallothionein isoform-specific responses and protein properties allow C. elegans to differentiate between toxic cadmium and essential zinc.