Role of metallothionein in cadmium traffic and toxicity in kidneys and other mammalian organs

Metallothioneins are cysteine-rich, small metal-binding proteins present in various mammalian tissues. Of the four common metallothioneins, MT-1 and MT-2 (MTs) are expressed in most tissues, MT-3 is predominantly present in brain, whereas MT-4 is restricted to the squamous epithelia. The expression of MT-1 and MT-2 in some organs exhibits sex, age, and strain differences, and inducibility with a variety of stimuli. In adult mammals, MTs have been localized largely in the cell cytoplasm, but also in lysosomes, mitochondria and nuclei. The major physiological functions of MTs include homeostasis of essential metals Zn and Cu, protection against cytotoxicity of Cd and other toxic metals, and scavenging free radicals generated in oxidative stress. The role of MTs in Cd-induced acute and chronic toxicity, particularly in liver and kidneys, is reviewed in more details. In acute toxicity, liver is the primary target, whereas in chronic toxicity, kidneys are major targets of Cd. The intracellular MTs bind Cd ions and form CdMT. In chronic intoxication, Cd stimulates de novo synthesis of MTs; it is assumed that toxicity in the cells starts when loading with Cd ions exceeds the buffering capacity of intracellular MTs. CdMT, released from the Cd-injured organs, or when applied parenterally for experimental purposes, reaches the kidneys via circulation, where it is filtered, endocytosed in the proximal tubule cells, and degraded in lysosomes. Liberated Cd can immediately affect the cell structures and functions. The resulting proteinuria and CdMT in the urine can be used as biomarkers of tubular injury.     

Antioxidant role of metallothioneins: a comparative overview.

Metallothioneins (MTs) are sulfhydryl-rich proteins binding essential and non-essential heavy metals. MTs display in vitro oxyradical scavenging capacity, suggesting that they may specifically neutralize hydroxyl radicals. Yet, this is probably an oversimplified view, as MTs represent a superfamily of widely differentiated metalloproteins. MT antioxidant properties mainly derive from sulfhydryl nucleophilicity, but also from metal complexation. Binding of transition metals displaying Fenton reactivity (Fe,Cu) can reduce oxidative stress, whereas their release exacerbates it. In vertebrates, MT gene promoters contain metal (MRE) and glucocorticoid response elements (GRE), Sp and AP sequences, but also antioxidant response elements (ARE). MT neosynthesis is induced by heavy metals, cytokines, hormones, but also by different oxidants and prooxidants. Accordingly, MT overexpression increases the resistance of tissues and cells to oxidative stress. As for invertebrates, data from the mussel show that MT can actually protect against oxidative stress, but is poorly inducible by oxidants. In yeast, there is a Cu(I)-MT that in contrast to mammalCu-MT exhibits antioxidant activity, possibly due to differences in metal binding domains. Finally, as the relevance of redox processes in cell signaling is becoming more and more evident, a search for MT effects on redox signaling could represent a turning point in the understanding of the functional role of these protein.     

Toxicity and interaction of titanium dioxide nanoparticles with microtubule protein

Titanium dioxide (TiO2) nanoparticles (NPs) are widely used in several manufactured products. The small size of NPs facilitates their uptake into cells as well as transcytosis across epithelial cells into blood and lymph circulation to reach different sites, such as the central nervous system. Different studies have shown the risks that TiO2 NPs in the neuronal system and other organs present. As membrane-bound layer aggregates or single particles, TiO2 NPs can enter not only cells, but also mitochondria and nuclei. Therefore these particles can interact with cytoplasmic proteins such as microtubules (MTs). MTs are cytoskeletal proteins that are essential in eukaryotic cells for a variety of functions, such as cellular transport, cell motility and mitosis. MTs in neurons are used to transport substances such as neurotransmitters. Single TiO2 NPs in cytoplasm can interact with these proteins and affect their crucial functions in different tissues. In this study, we showed the effects of TiO2 NPs on MT polymerization and structure using ultraviolet spectrophotometer and fluorometry. The fluorescent spectroscopy showed a significant tubulin conformational change in the presence of TiO2 NPs and the ultraviolet spectroscopy results showed that TiO2 NPs affect tubulin polymerization and decrease it. The aim of this study was to find the potential risks that TiO2 NPs pose to human organs and cells.     

无脊椎动物金属硫蛋白(MTs)多样性及其生态服务功能

金属硫蛋白（MTs）是一类低分子量、半胱氨酸含量异常丰富的金属结合多肽,自从20世纪70年代中期发现海洋无脊椎动物MTs以来,MTs已被证明广泛存在于无脊椎动物的各个类群之中。无脊椎动物物种间的金属硫蛋白存在着显著差异,研究无脊椎动物MTs多样性并揭示其生态服务功能,在理论与实践上都至关重要。本文分析了无脊椎动物MTs的多样性：结合金属元素多样性、同形体及其变体的蛋白质遗传多样性和生态服务功能多样性,并讨论了 MTs的三个生态服务功能：MTs对重金属解毒和调节作用、MTs作为环境监测的生物标志物、MTs的环境重金属污染净化功能及其在环境污染治理中的作用。     

Metallothionein induction and heavy metal accumulation in white shrimp Litopenaeus vannamei exposed to cadmium and zinc

Metallothioneins (MTs) have been widely considered for their potential use as specific biomarkers to reflect the existence of heavy metal pollution, because their induction has been observed to be obviously elevated after heavy metal exposure in a large number organism studied. However, relatively fewer efforts have been made in MT-related studies of prawn species, such as the white shrimp Litopenaeus vannamei, a globally important aquaculture species. With the results from gel filtration chromatography, we demonstrate the existence of MTs or MT-like proteins in L.vannamei. We further studied the relationship between MT induction and metals accumulation after long-term exposure to the heavy metals Cd and Zn. From our results, it is very clear that the response of L. vannamei to Cd differs from that to Zn, and this should be considered when using MTs in field applications to monitor metals contamination.     

Induction, regulation, degradation, and biological significance of mammalian metallothioneins

MTs are small cysteine-rich metal-binding proteins found in many species and, although there are differences between them, it is of note that they have a great deal of sequence and structural homology. Mammalian MTs are 61 or 62 amino acid polypeptides containing 20 conserved cysteine residues that underpin the binding of metals. The existence of MT across species is indicative of its biological demand, while the conservation of cysteines indicates that these are undoubtedly central to the function of this protein. Four MT isoforms have been found so far, MT-1, MT-2, MT-3, and MT-4, but these also have subtypes with 17 MT genes identified in man, of which 10 are known to be functional. Different cells express different MT isoforms with varying levels of expression perhaps as a result of the different function of each isoform. Even different metals induce and bind to MTs to different extents. Over 40 years of research into MT have yielded much information on this protein, but have failed to assign to it a definitive biological role. The fact that multiple MT isoforms exist, and the great variety of substances and agents that act as inducers, further complicates the search for the biological role of MTs. This article reviews the current knowledge on the biochemistry, induction, regulation, and degradation of this protein in mammals, with a particular emphasis on human MTs. It also considers the possible biological roles of this protein, which include participation in cell proliferation and apoptosis, homeostasis of essential metals, cellular free radical scavenging, and metal detoxification.     

Induction,Regulation, Degradation,and Biological Significance of Mammalian Metallothioneins

MTs are small cysteine-rich metal-binding proteins found in many species and, although there are differences between them, it is of note that they have a great deal of sequence and structural homology. Mammalian MTs are 61 or 62 amino acid polypep-tides containing 20 conserved cysteine residues that underpin the binding of metals. The existence of MT across species is indicative of its biological demand, while the conservation of cysteines indicates that these are undoubtedly central to the function of this protein. Four MT isoforms have been found so far, MT-1, MT-2, MT-3, and MT-4, but these also have subtypes with 17 MT genes identified in man, of which 10 are known to be functional. Different cells express different MT isoforms with varying levels of expression perhaps as a result of the different function of each isoform. Even different metals induce and bind to MTs to different extents. Over 40 years of research into MT have yielded much information on this protein, but have failed to assign to it a definitive biological role. The fact that multiple MT isoforms exist, and the great variety of substances and agents that act as inducers, further complicates the search for the biological role of MTs. This article reviews the current knowledge on the biochemistry, induction, regulation, and degradation of this protein in mammals, with a particular emphasis on human MTs. It also considers the possible biological roles of this protein, which include participation in cell proliferation and apoptosis, homeostasis of essential metals, cellular free radical scavenging, and metal detoxification.     

Metallothioneins in liver of Rutilus rutilus exposed to Cu2+. Analysis by metal summation, SH determination and spectrofluorimetry

Metallothioneins (MTs) have important roles in the homeostasis of essential metals and in the detoxication of heavy metals. They also represent a potential indicator of aquatic contamination by metals. Routine methods are needed for MTs quantification in ecotoxicological studies. This paper investigates the possibility to use the spectrofluorescent properties of Cu-MTs for MTs quantification. Cu displacement of metals coordinated to MTs and spectrofluorimetric determination of the obtained Cu-MTs was tested with commercial MTs and Cu2+-induced MTs in roach liver (Rutilus rutilus). Results of this original and simple spectrofluorimetric quantification of MTs presented a good correlation with data obtained with SH quantification, but not with metal summation evaluation of MTs (analysis of Zn, Cu and Cd coordinated to MTs). The three methods showed an clear induction of MTs in roach liver after 7 days of Cu2+ exposure. After 14 days of contamination, a reduction of hepatic MTs content was observable and not correlated to liver recovery. Results show that this low cost spectrofluorimetric method is useful to quantify MTs.     

Metallothionein induction in human peripheral blood lymphocytes by heavy metals

Human peripheral blood lymphocytes have the capacity to produce metallothioneins (MTs) as a protective response to cadmium exposure. To define the range of metal species inducing lymphocyte MTs, cellular proteins synthesized after exposure to each of 11 heavy metals were analyzed by gel electrophoresis. Toxic metals such as cadmium, mercury and silver were found to induce thioneins (apoproteins of MTs) at relatively low concentrations (maximum at approximately 10 microM), whereas less toxic metals such as zinc, copper and nickel were inductive at relatively high concentrations (maximum at approximately 200 microM). Tin, lead, iron, cobalt, and manganese did not induce thioneins. The heavy metal specificity of MT induction in the lymphocyte resembles that in the liver, and the regulatory mechanism of MT production seems to be similar in both of these tissues. In the cells exposed to highly toxic metals such as cadmium and mercury, expression of cytotoxicity (represented by decline of cysteine uptake) was remarkable at the metal concentrations higher than those saturating thionein induction, supporting the protective role of MTs against heavy metals.     

From heavy metal‐binders to biosensors: Ciliate metallothioneins discussed

Metallothioneins (MTs) are ubiquitous proteins with the capacity to bind heavy metal ions (mainly Cd, Zn or Cu), and they have been found in animals, plants, eukaryotic and prokaryotic micro‐organisms. We have carried out a comparative analysis of ciliate MTs (Tetrahymena species) to well‐known MTs from other organisms, discussing their exclusive features, such as the presence of aromatic amino acid residues and almost exclusive cysteine clusters (CCC) present in cadmium‐binding metallothioneins (CdMTs), higher heavy metal‐MT stoichiometry values, and a strictly conserved modular–submodular structure. Based on this last feature and an extensive gene duplication, we propose a possible model for the evolutionary history of T. thermophila MTs. We also suggest possible functions for these MTs from consideration of their differential gene expressions and discuss the potential use of these proteins and/or their gene promoters for designing molecular or whole‐cell biosensors for a fast detection of heavy metals in diverse polluted ecosystems.     

Hsp90 binds microtubules and is involved in the reorganization of the microtubular network in angiosperms

Microtubules (MTs) are essential for many processes in plant cells. MT-associated proteins (MAPs) influence MT polymerization dynamics and enable them to perform their functions. The molecular chaperone Hsp90 has been shown to associate with MTs in animal and plant cells. However, the role of Hsp90-MT binding in plants has not yet been investigated. Here, we show that Hsp90 associates with cortical MTs in tobacco cells and decorates MTs in the phragmoplast. Further, we show that tobacco Hsp90_MT binds directly to polymerized MTs in vitro. The inhibition of Hsp90 by geldanamycin (GDA) severely impairs MT re-assembly after cold-induced de-polymerization. Our results indicate that the plant Hsp90 interaction with MTs plays a key role in cellular events, where MT re-organization is needed.     

Proteomic analysis of proteins secreted by Botrytis cinerea in response to heavy metal toxicity

Although essential in many cellular processes, metals become toxic when they are present in excess and constitute a global environmental hazard. To overcome this stress, fungi have evolved several mechanisms at both intracellular and extracellular levels. In particular, fungi are well known for their ability to secrete a large panel of proteins. However, their role in the adaptation of fungi to metal toxicity has not yet been investigated. To address this question, here, the fungus Botrytis cinerea was challenged to copper, zinc, nickel or cadmium stress and secreted proteins were collected and separated by 2D-PAGE. One hundred and sixteen spots whose volume varied under at least one tested condition were observed on 2D gels. Densitometric analyses revealed that the secretome signature in response to cadmium was significantly different from those obtained with the other metals. Fifty-five of these 116 spots were associated with unique proteins and functional classification revealed that the production of oxidoreductases and cell-wall degrading enzymes was modified in response to metals. Promoter analysis disclosed that PacC/Rim101 sites were statistically over-represented in the upstream sequences of the 31 genes corresponding to the varying unique spots suggesting a possible link between pH regulation and metal response in B. cinerea.     

Cellular mechanisms for heavy metal detoxification and tolerance.

Heavy metals such as Cu and Zn are essential for normal plant growth, although elevated concentrations of both essential and non-essential metals can result in growth inhibition and toxicity symptoms. Plants possess a range of potential cellular mechanisms that may be involved in the detoxification of heavy metals and thus tolerance to metal stress. These include roles for the following: for mycorrhiza and for binding to cell wall and extracellular exudates; for reduced uptake or efflux pumping of metals at the plasma membrane; for chelation of metals in the cytosol by peptides such as phytochelatins; for the repair of stress-damaged proteins; and for the compartmentation of metals in the vacuole by tonoplast-located transporters. This review provides a broad overview of the evidence for an involvement of each mechanism in heavy metal detoxification and tolerance.     

Cysteine Cathepsins and the Cutting Edge of Cancer Invasion

The cellular response to heat stress includes the induction of a group of proteins called the Heat Shock Proteins, whose functions include the synthesis of the thermoprotectant trehalose, refolding of denatured proteins, and ubiquitin- and proteasome-dependent degradation. Recent studies show that simply increasing the activity of ubiquitin- and proteasome-dependent degradation can replace the essential functions played by the induction of heat shock proteins during a heat stress. These results suggest that accumulation of denatured or aggregated proteins is the reason for the loss of cell viability due to heat stress.     

Insights into environmental bioremediation by microorganisms through functional genomics and proteomics

Environmental pollutants in the soil are a major concern worldwide. Bioremediation mediated by microorganisms is a highly promising technology that is environmentally friendly, safe, and effective. However, incomplete biological information regarding the cellular responses in many microbial communities restricts progress in the site-specific mineralization process. The application of proteomics in environmental bioremediation research provides a global view of the protein compositions of the microbial cells and offers a promising approach to address the molecular mechanisms of bioremediation. With the combination of proteomics, functional genomics provide an insight into global metabolic and regulatory networks that can enhance the understanding of gene functions. This article deals with the applications of functional genomics and proteomics to dissect the cellular responses to environmental stimuli, such as stress response, induction and expressions of regulatory proteins/enzymes in response to aromatic hydrocarbons and heavy metals. An understanding of the growth conditions governing the expression of the proteome (for example, enzymes and regulatory proteins of aromatic hydrocarbon degradation, energy generation pathways, transport and stress-related proteins) in a specific environment is essential for developing rational strategies for successful bioremediation.     

高等植物重金属耐性与超积累特性及其分子机理研究

由于重金属污染日益严重, 重金属在土壤_植物系统中的行为引起了人们的高度重视。高等植物对重金 属的耐性与积累性, 已经成为污染生态学研究的热点。近年来, 由于分子生态学等学科的发展, 有关植物对重金属的解毒和耐性机理、重金属离子富集机制的研究取得了较大进展。高等植物对重金属的耐性和积累在种间和基因型之间存在很大差异。根系是重金 属等土壤污染物进入植物的门户。根系分泌物改变重金属的生物有效性和毒性, 并在植物吸收重金属的过程中发挥重要作用。土壤中的大部分重金属离子都是通过金属转运蛋白进入根细胞, 并在植物体内进一步转运至液泡贮存。在重金属胁迫条件下植物螯合肽 (PC) 的合成是植物对胁迫的一种适应性反应。耐性基因型合成较多的PC, 谷胱甘肽 (GSH) 是合成PC的前体, 重金属与PC螯合并转移至液泡中贮存, 从而达到解毒效果。金属硫蛋白 (MTs) 与PC一样, 可以与重金属离子螯合, 从而降低重金属离子的毒性。该文从分子水平上论述了根系分泌物、金属转运蛋白、MTs、PC、GSH在重金属耐性及超积累性中的作用, 评述了近 10年来这方面的研究进展, 并在此基础上提出存在的问题和今后研究的重点。