Loss of the metal binding properties of metallothionein induced by hydrogen peroxide and free radicals

Jiménez, I.; Gotteland, Martín; Zarzuelo, Antonio; Uauy, Ricardo; Speisky, Hernán

doi:10.1016/s0300-483x(97)03638-x

Cited by 50 publications

(25 citation statements)

References 20 publications

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“…MT-I has a wide tissue distribution, with its primary role being the detoxification of cadmium and maintenance zinc homeostasis. However, MT-1 has also been shown to be a potent oxygen radical scavenger [19][20][21][22][23] and thus can protect cells from oxidative stress. Furthermore, MTs have been shown to protect DNA from oxidative damage, 24 and to protect cells from apoptosis induced by some exogenous stimuli.…”

Section: Discussionmentioning

confidence: 99%

Cellular response to conditional expression of hepatitis C virus core protein in Huh7 cultured human hepatoma cells

Prow

Lemon

et al. 2002

Hepatology

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Data suggesting that the hepatitis C virus (HCV) core protein influences normal cellular processes remain controversial. To determine the effects of core on cellular gene expression in hepatocytes, we developed a human hepatoma (Huh7)-derived cell line with tightly regulated core expression under the control of a tetracycline-regulated promoter. Cells expressing core did not have impaired proliferative abilities. Changes in gene expression profiles in response to core expression were determined using commercial oligonucleotide microarrays (Affymetrix GeneChip). Significant increases were observed in the abundance of mRNA-encoding members of the metallothionein (MT) family, as well as nicotinamide N-methyltransferase (NNMT) and glutathione peroxidase-like protein (GPLP). These changes did not result from removal of tetracycline from growth media, and were confirmed in reverse-transcription polymerase chain reaction (RT-PCR) assays. They suggest that core protein expression leads to intracellular oxidative stress, and that vital cellular functions are, in turn, protected by up-regulation of cellular antioxidant defense mechanisms. In conclusion, these findings can explain many potentially conflicting prior observations concerning the effects of core on cellular physiology, and are of relevance to the role of core protein in the pathogenesis of HCV-related fibrosis and hepatocellular carcinoma. (HEPATOLOGY 2002;35:1237-1246

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Section: Discussionmentioning

confidence: 99%

Cellular response to conditional expression of hepatitis C virus core protein in Huh7 cultured human hepatoma cells

Prow

Lemon

et al. 2002

Hepatology

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“…Furthermore, an elegant study by Pearce et al. (14) has shown that green fluorescent protein-modified MT undergoes conformational changes in the presence of NO in cultured pulmonary artery endothelial cells leading to the release of metals from MT.MT becomes oxidized under conditions of oxidative and nitrosative stress in vitro (11,13,(15)(16)(17)(18). MT nitrosylation leads to disulfide bond formation in the ␤-domain (11).…”

mentioning

confidence: 99%

“…MT becomes oxidized under conditions of oxidative and nitrosative stress in vitro (11,13,(15)(16)(17)(18). MT nitrosylation leads to disulfide bond formation in the ␤-domain (11).…”

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confidence: 99%

Metallothionein Disulfides Are Present in Metallothionein-overexpressing Transgenic Mouse Heart and Increase under Conditions of Oxidative Stress

Feng¹,

Benz

Cai

et al. 2006

Journal of Biological Chemistry

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Metallothionein (MT) releases zinc under oxidative stress conditions in cultured cells. The change in the MT molecule after zinc release in vivo is unknown although in vitro studies have identified MT disulfide bond formation. The present study was undertaken to test the hypothesis that MT disulfide bond formation occurs in vivo. A cardiac-specific MT-overexpressing transgenic mouse model was used. Mice were administered saline as a control or doxorubicin (20 mg/kg), which is an effective anticancer drug but with severe cardiac toxicity at least partially because of the generation of reactive oxygen species. A differential alkylation of cysteine residues in MT of the heart extracts was performed. Free and metal-bound cysteines were first trapped by N-ethylmaleimide and the disulfide bonds were reduced by dithiothreitol followed by alkylation with radiolabeled iodoacetamide. Analyses of the differentially alkylated MTs in the heart extract by high preformance liquid chromatography, SDS-PAGE, Western blot, and mass spectrometry revealed that disulfide bonds were present in MT in vivo under both physiological and oxidative stress conditions. More disulfide bonds were found in MT under the oxidative stress conditions. The MT disulfide bonds were likely intramolecular and both ␣-and ␤-domains were involved in the disulfide bond formation, although the ␣-domain appeared to be more easily oxidized than the ␤-domain. The results suggest that under physiological conditions, the formation of MT disulfide bonds is involved in the regulation of zinc homeostasis. Additional zinc release from MT under oxidative stress conditions is accompanied by more MT disulfide bond formation. Previous studies have shown that metallothionein (MT)2 functions in cardiac protection against oxidative damage (1-4). The mechanism of the antioxidant action of MT remains elusive. It has been shown in vitro that MT directly interacts with reactive oxygen and nitrogen species to prevent oxidative and nitrosative damage to macromolecules such as lipids, proteins, and DNA (5-8). However, this interaction has never been demonstrated in vivo and would unlikely be a major mechanism for the antioxidant action of MT in vivo because the reactivity of reactive oxygen and nitrogen species would only allow their reaction with molecules that have a close proximity to their generation sites. Alternatively, in vitro studies have demonstrated that the multiple cysteine residues of MT can be oxidized. The sulfur clusters that bind zinc in MT create an oxidoreductive environment for zinc at a redox potential so low that MT can be readily oxidized by mild cellular oxidants, such as glutathione disulfide, with the release of zinc (9). This MT oxidation and zinc release process would play a critical role in the cellular response to oxidative and nitrosative stress.The high content of cysteine residues enables MT to avidly bind zinc and other metals (10). The binding of divalent metals to MT occurs in two dumbbell-shaped domains, of which the N-terminal ␤-domain usually...

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“…Biochemical studies have shown that the particularly high affinity of metallothioneins for cadmium and copper and the oxidation of their sulfhydryl groups by reaction products of hydrogen peroxide can liberate zinc (31,54,63,71). Moreover, metallothionein genes are well-defined targets of MTF-1, i.e., their transcription is induced by MTF-1 in response to heavy metals.…”

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Activity of Metal-Responsive Transcription Factor 1 by Toxic Heavy Metals and H₂O₂ In Vitro Is Modulated by Metallothionein

Zhang

Georgiev

Hagmann

et al. 2003

Molecular and Cellular Biology

219

152

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Metallothioneins are small, cysteine-rich proteins that avidly bind heavy metals such as zinc, copper, and cadmium to reduce their concentration to a physiological or nontoxic level. Metallothionein gene transcription is induced by several stimuli, notably heavy metal load and oxidative stress. Transcriptional induction of metallothionein genes is mediated by the metal-responsive transcription factor 1 (MTF-1), an essential zinc finger protein that binds to specific DNA motifs termed metal-response elements. In cell-free DNA binding reactions with nuclear extracts, MTF-1 requires elevated zinc concentrations for efficient DNA binding but paradoxically is inactivated by other in vivo inducers such as cadmium, copper, and hydrogen peroxide. Here we have developed a cell-free, MTF-1-dependent transcription system which accurately reproduces the activation of metallothionein gene promoters not only by zinc but also by these other inducers. We found that while transcriptional induction by zinc can be achieved by elevated zinc concentration alone, induction by cadmium, copper, or H 2 O 2 additionally requires the presence of zinc-saturated metallothionein. This is explained by the preferential binding of cadmium or copper to metallothionein or its oxidation by H 2 O 2 ; the concomitant release of zinc in turn leads to the activation of transcription factor MTF-1. Conversely, thionein, the metal-free form of metallothionein, inhibits activation of MTF-1. The release of zinc from cellular components, including metallothioneins, and the sequestration of zinc by newly produced apometallothionein might be a basic mechanism to regulate MTF-1 activity upon cellular stress.All living organisms are able to cope with a variety of stress situations by immediately adapting their gene expression program to the stress stimulus. For example, metallothioneins, small cysteine-rich proteins, are strongly upregulated upon heavy metal load (2,27,29,49,63,70,71). They have the ability to bind and hence neutralize toxic (such as Cd and Hg) and excess nontoxic (such as Zn and Cu) heavy metal ions and also act as radical scavengers. Metal response element binding transcription factor 1, also called metal-responsive transcription factor 1 (MTF-1), plays an important role in the cellular response to heavy metal stress (2,21,38,55,75) and is essential for embryonic liver development in the mouse (24). MTF-1 contains six zinc fingers of the C 2 H 2 type. C-terminal to the zinc fingers are three distinct activation domains, an acidic, a proline-rich, and a serine/threonine-rich domain. Via the zinc fingers, it binds to DNA sequence motifs with the consensus binding site TGCRCNC, known as metal response elements (MREs). MREs are present in the promoters of metallothionein genes (41,61,64) and other target genes, including zinc transporter ZnT1 and ␥-glutamylcysteine synthetase heavy chain (24,34,37). MTF-1 is also involved in the responses to oxidative stress (16, 24), hypoxia (23, 46), and amino acid starvation (1). In quiescent cells, MTF-1 pref...

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Loss of the metal binding properties of metallothionein induced by hydrogen peroxide and free radicals

Cited by 50 publications

References 20 publications

Cellular response to conditional expression of hepatitis C virus core protein in Huh7 cultured human hepatoma cells

Cellular response to conditional expression of hepatitis C virus core protein in Huh7 cultured human hepatoma cells

Metallothionein Disulfides Are Present in Metallothionein-overexpressing Transgenic Mouse Heart and Increase under Conditions of Oxidative Stress

Activity of Metal-Responsive Transcription Factor 1 by Toxic Heavy Metals and H₂O₂ In Vitro Is Modulated by Metallothionein

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Loss of the metal binding properties of metallothionein induced by hydrogen peroxide and free radicals

Cited by 50 publications

References 20 publications

Cellular response to conditional expression of hepatitis C virus core protein in Huh7 cultured human hepatoma cells

Cellular response to conditional expression of hepatitis C virus core protein in Huh7 cultured human hepatoma cells

Metallothionein Disulfides Are Present in Metallothionein-overexpressing Transgenic Mouse Heart and Increase under Conditions of Oxidative Stress

Activity of Metal-Responsive Transcription Factor 1 by Toxic Heavy Metals and H2O2 In Vitro Is Modulated by Metallothionein

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Activity of Metal-Responsive Transcription Factor 1 by Toxic Heavy Metals and H₂O₂ In Vitro Is Modulated by Metallothionein