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![Metallothionein (MT) structure. Model of two binding sites of metallothionein. Red big beads are metal atoms (e.g., Zn), and small yellow beads are sulfur atoms. Adopted and modified according to [27].](publication/236055564/figure/fig1/AS:299359037083654@1448384164463/Metallothionein-MT-structure-Model-of-two-binding-sites-of-metallothionein-Red-big.png)
Metallothionein (MT) structure. Model of two binding sites of metallothionein. Red big beads are metal atoms (e.g., Zn), and small yellow beads are sulfur atoms. Adopted and modified according to [27].
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![Table 1 . Summary of reactive oxygen and nitrogen species [10,11].](publication/236055564/figure/tbl1/AS:670524560646157@1536876921566/Summary-of-reactive-oxygen-and-nitrogen-species-10-11_Q320.jpg)
Free radicals are chemical particles containing one or more unpaired electrons, which may be part of the molecule. They cause the molecule to become highly reactive. The free radicals are also known to play a dual role in biological systems, as they can be either beneficial or harmful for living systems. It is clear that there are numerous mechanis...
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Context 1
... these low molecular weight cysteine-rich proteins have been continuously studied in all aspects, including physical, chemical and biochemical properties. Mammalian MTs may contain 61–68 amino acids, and among them 20 are cysteines [16,17]. These unique proteins are involved in diverse intracellular functions [18], but their role in the detoxification of heavy metals and in the maintaining of essential metal ion homeostasis, which is due to their high affinity for these metals, is mostly investigated [19,20]. For mammals, MTs bind zinc [21], but with excess copper or cadmium, zinc can be easily replaced by these metals [22]. Cells that contain excessive amounts of MTs are resistant to cadmium toxicity [23], while cell lines that cannot synthesize MTs are sensitive to cadmium [24]. Genetic studies using transgenic or knockout mouse models are further evidence of the role of MTs in protection against cadmium toxicity [25,26]. Based on structural models, it can be assumed that the MT molecule is composed of two binding domains, α and β , which are composed of cysteine clusters. Covalent binding of metal atoms involves sulfhydryl cysteine residues ( Figure 1). The N -terminal part of the peptide is designated as β -domain and has three binding sites for divalent ions, and the C-terminal part (the α -domain) has the ability to bind four divalent metal ions. Four mammalian MT isoforms (MT-1–MT-4) and 13 MT-like human proteins were identified [28]. The differences of constituent forms come mainly from post-translational modifications, small changes in primary structure, type of incorporated metal ion and speed of degradation. Despite the physical-chemical similarity of the forms, their roles and occurrence in tissues vary significantly [29]. MT-1 and MT-2 are present almost in all types of soft tissues [30–32], MT-3 is expressed mostly in brain tissue, but also in heart, kidneys and reproductive organs [33,34] and the MT-4 gene was detected in stratified squamous epithelial cells associated with oral epithelia, esophagus, upper stomach, tail, footpads and neonatal skin. [35]. In humans, the MT genes are located on chromosome 16 in a cluster and involve 16 identified genes, from which five are pseudogenes [36]. Although the MT-II, MT-III and MT-IV proteins are encoded by a single gene, the MT-I protein comprises ...
Context 2
... on structural models, it can be assumed that the MT molecule is composed of two binding domains, α and β, which are composed of cysteine clusters. Covalent binding of metal atoms involves sulfhydryl cysteine residues (Figure 1). The N-terminal part of the peptide is designated as β-domain and has three binding sites for divalent ions, and the C-terminal part (the α-domain) has the ability to bind four divalent metal ions. ...
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Citations
... Metallothioneins are a family of small (~6 kDa) proteins containing stretches of 20 cysteine residues, which allow them to bind ions of heavy metals (gold, silver, cadmium, etc.) [54,55]. Metallothioneins have been found in different organisms [56]; they are involved in protection against the toxic effects of heavy metal salts [57] and oxidative stress [58]. ...
Electron microscopy (EM) is one of the most efficient methods for studying the fine structure of cells with a resolution thousands of times higher than that of visible light microscopy. The most advanced implementation of electron microscopy in biology is EM tomography of samples stabilized by freezing without water crystallization (cryoET). By circumventing the drawbacks of chemical fixation and dehydration, this technique allows investigating cellular structures in three dimensions at the molecular level, down to resolving individual proteins and their subdomains. However, the problem of efficient identification and localization of objects of interest has not yet been solved, thus limiting the range of targets to easily recognizable or abundant subcellular components. Labeling techniques provide the only way for locating the subject of investigation in microscopic images. CryoET imposes conflicting demands on the labeling system, including the need to introduce into a living cell the particles composed of substances foreign to the cellular chemistry that have to bind to the molecule of interest without disrupting its vital functions and physiology of the cell. This review examines both established and prospective methods for selective labeling of proteins and subcellular structures aimed to enable their localization in cryoET images.
... We also detected increased expression of sirtuin 2 (Sirt2), a fatty-acid oxidation sensitive enzyme that regulates NAD+ availability, for treated MuSCs when compared to controls ( Fig. 4H) (45,46). Conversely, upregulated genes in untreated cells included metallothioneins 1 and 2 (Mt1, Mt2), markers of oxidative stress, and Rock2, a negative regulator of mitophagy (47,48). GO Term enrichment analysis showed that treatment reduced apoptotic pathways, cell cycle entry, and TOR signaling (Supp. ...
Adult stem cells decline in number and function in old age and identifying factors that can delay or revert age-associated adult stem cell dysfunction are vital for maintaining healthy lifespan. Here we show that Vitamin A, a micronutrient that is derived from diet and metabolized into retinoic acid, acts as an antioxidant and transcriptional regulator in muscle stem cells. We first show that obstruction of dietary Vitamin A in young animals drives mitochondrial and cell cycle dysfunction in muscle stem cells that mimics old age. Next, we pharmacologically targeted retinoic acid signaling in myoblasts and aged muscle stem cells ex vivo and in vivo and observed reductions in oxidative damage, enhanced mitochondrial function, and improved maintenance of quiescence through fatty acid oxidation. We next detected the receptor for vitamin A derived retinol, stimulated by retinoic acid 6 or Stra6, was diminished with muscle stem cell activation and in old age. To understand the relevance of Stra6 loss, we knocked down Stra6 and observed an accumulation of mitochondrial reactive oxygen species, as well as changes in mitochondrial morphology and respiration. These results demonstrate that Vitamin A regulates mitochondria and metabolism in muscle stem cells and highlight a unique mechanism connecting stem cell function with vitamin intake.
... It is known that their reactivity can be significantly changed in the presence of vitamins C and E [36]. Selenium compounds can reduce peroxides and have a great influence on the release of zinc by metallothionein [22,34,37]. However, the lowest OS value was calculated for the sample with BiVits® ACTIVA Recovery, which possesses the strongest antioxidant effect. ...
... In addition, due to their high thiol content, these proteins were involved in quenching hydroxyl (OH . ) and superoxide radicals (O 2 -. ) [107]. Under our experimental conditions, the significant increase of MT levels in PEN-exposed clams reflected probably an adaptive approach adopted by R. decussates to reduce this fungicide. ...
The baleful effect of penconazole fungicide on clam Ruditapes decussatus gills: fatty acids composition, redox status and histological features, Chemistry and Ecology, ABSTRACT Penconazole is a widely used fungicide to control critical fungal diseases of crops. This study aimed to elucidate the effects of PEN exposure, for 96 h, on fatty acids composition, redox status, and histopathological injuries in Ruditapes decussatus gills under different concentrations (4, 40, and 400 µg/L). Compared to the control group, our results showed an increase in saturated and monounsaturated fatty acids. However, the levels of polyunsaturated fatty acids were diminished, mainly those of eicosapentaenoic, docosahexaenoic, and arachidonic acids upon exposure to PEN. Depletion of PUFAs' levels, particularly the long-chain and double-bond-rich ones, can affect membrane fluidity, and disrupt membranes' biological functions, such as cell signalling, membrane transport, and protection against other environmental stresses. Herein, PEN uptake also results in enhanced oxidative stress by increasing levels of the ferric-reducing antioxidant power, hydrogen peroxidase, malondialdehyde, advanced oxidation protein products, and protein carbonyls. Moreover, changes in both enzymatic and non-enzymatic antioxidants were noticeable, indicating the stimulation of the antioxidant defense system. Adverse histological alterations were detected in PEN-treated specimens' gill tissues, supporting our biochemical findings. Overall, in the current study, redox status was related to changes in fatty acid composition, contributing to a better understanding of the PEN-toxicity mechanism in clams.
... The main difference in elemental composition between diesel PM and brake-wear PM measured here is the enrichment of metals in the latter. Consistent with this difference, we demonstrated that all brake-wear PM types could upregulate the expression of genes encoding metallothioneins, cysteine-rich metal-chelating proteins [49]. Metallothioneins are canonically described as zinc-binding proteins [50], which play an important role in regulating the intracellular free concentration of zinc [49]. ...
... Consistent with this difference, we demonstrated that all brake-wear PM types could upregulate the expression of genes encoding metallothioneins, cysteine-rich metal-chelating proteins [49]. Metallothioneins are canonically described as zinc-binding proteins [50], which play an important role in regulating the intracellular free concentration of zinc [49]. When intracellular free zinc concentrations increase, zinc binds to metal regulatory transcription factor 1 (MTF1), promoting nuclear translocation and binding to and Oxidative Stress Score. ...
... In C and D, a Pearson's correlation was used. In F and G, and two-tailed paired t-test was used promoter sequences on metallothionein genes, increasing their expression to maintain zinc homeostasis [49]. In this context, it is notable that SemiMxCu-derived brakewear PM contained more zinc than any of the other vehicle-derived PM types examined, however, NAO and ceramic brake-wear PM still induced the highest magnitude response in metallothionein gene expression. ...
Background
Airborne fine particulate matter with diameter < 2.5 μm (PM2.5), can reach the alveolar regions of the lungs, and is associated with over 4 million premature deaths per year worldwide. However, the source-specific consequences of PM2.5 exposure remain poorly understood. A major, but unregulated source is car brake wear, which exhaust emission reduction measures have not diminished.
Methods
We used an interdisciplinary approach to investigate the consequences of brake-wear PM2.5 exposure upon lung alveolar cellular homeostasis using diesel exhaust PM as a comparator. This involved RNA-Seq to analyse global transcriptomic changes, metabolic analyses to investigate glycolytic reprogramming, mass spectrometry to determine PM composition, and reporter assays to provide mechanistic insight into differential effects.
Results
We identified brake-wear PM from copper-enriched non-asbestos organic, and ceramic brake pads as inducing the greatest oxidative stress, inflammation, and pseudohypoxic HIF activation (a pathway implicated in diseases associated with air pollution exposure, including cancer, and pulmonary fibrosis), as well as perturbation of metabolism, and metal homeostasis compared with brake wear PM from low- or semi-metallic pads, and also, importantly, diesel exhaust PM. Compositional and metal chelator analyses identified that differential effects were driven by copper.
Conclusions
We demonstrate here that brake-wear PM may perturb cellular homeostasis more than diesel exhaust PM. Our findings demonstrate the potential differences in effects, not only for non-exhaust vs exhaust PM, but also amongst different sources of non-exhaust PM. This has implications for our understanding of the potential health effects of road vehicle-associated PM. More broadly, our findings illustrate the importance of PM composition on potential health effects, highlighting the need for targeted legislation to protect public health.
... More specifically, genes encoding Metallothioneins (MT1E, MT2A, and MT1X) were among the top six genes that significantly upregulated following FOS upregulation (FDR < 1e-50) ( Fig. 3i and Supplementary Data 1). Metallothioneins have been wellknown for their antioxidant and antiapoptotic effects, and their role in T2D has gained increasing attention in recent years [34][35][36] . A previous study also reported the function of FOS and JUND in preventing cell death from oxidative stress 37 , consistent with regX's interpretations (Table 1 and Fig. 3f, g). ...
Cells are regulated at multiple levels, from regulations of individual genes to interactions across multiple genes. Some recent neural network models can connect molecular changes to cellular phenotypes, but their design lacks modeling of regulatory mechanisms, limiting the decoding of regulations behind key cellular events, such as cell state transitions. Here, we present regX, a deep neural network incorporating both gene-level regulation and gene-gene interaction mechanisms, which enables prioritizing potential driver regulators of cell state transitions and providing mechanistic interpretations. Applied to single-cell multi-omics data on type 2 diabetes and hair follicle development, regX reliably prioritizes key transcription factors and candidate cis-regulatory elements that drive cell state transitions. Some regulators reveal potential new therapeutic targets, drug repurposing possibilities, and putative causal single nucleotide polymorphisms. This method to analyze single-cell multi-omics data demonstrates how the interpretable design of neural networks can better decode biological systems.
... For instance, z plays a key role in antioxidant defense systems, DNA repair, and enzyme functio However, Cd disrupts Zn homeostasis in astrocytes by competing with Zn for sites, thereby displacing Zn and perturbing its normal functions [204,205]. This eve weakens the antioxidant defense of glial cells and causes CNS disease susceptibilit Also, considering that Zn is responsible for the functioning of metallothioneins (a that catalyzes the detoxification of metals against oxidative stress), the Cd-indu homeostasis imbalance can further lead to uncontrolled cellular damage that is imp in many neurodegenerative diseases [207,208]. ...
... This eventually weakens the antioxidant defense of glial cells and causes CNS disease susceptibility [206]. Also, considering that Zn is responsible for the functioning of metallothioneins (a protein that catalyzes the detoxification of metals against oxidative stress), the Cd-induced Zn homeostasis imbalance can further lead to uncontrolled cellular damage that is implicated in many neurodegenerative diseases [207,208]. ...
Citation: Ijomone, O.K.; Ukwubile, I.I.; Aneke, V.O.; Olajide, T.S.; Inyang, H.O.; Omotosho, O.I.; Oyerinde, T.O.; Anadu, V.E.; Gbayisomore, T.J.; Okeowo, O.M.; et al. Glial Perturbation in Metal Neurotoxicity: Implications for Brain Disorders. Neuroglia 2025, 6, 4. https://doi. Abstract: Overexposure of humans to heavy metals and essential metals poses a significant risk for the development of neurological and neurodevelopmental disorders. The mechanisms through which these metals exert their effects include the generation of reactive oxygen species, mitochondrial dysfunction, activation of inflammatory pathways, and disruption of cellular signaling. The function of glial cells in brain development and in the maintenance of homeostasis cannot be overlooked. The glial cells are particularly susceptible to metal-induced neurotoxicity. Accumulation of metals in the brain promotes microglial activation, triggering inflammatory responses that can coincide with other mechanisms of neurotoxicity, inducing alteration in synaptic transmission, cognitive deficit, and neuronal damage. In this review, we highlighted the role of glial dysfunction in some selected neurodegenerative diseases and neurodevelopmental disorders. We further dive into how exposure to metals such as nickel, manganese, methyl mercury, cadmium, iron, arsenic, and lead affect the functions of the microglia, astrocytes, and oligodendrocytes and the mechanisms through which they exert the effects on the brain in relation to some selected neurodegenerative diseases and neurodevelopmental disorders. Potential therapeutic interventions such as the use of new and improved chelating agents and antioxidant therapies might be a significant approach to alleviating these metal-induced glial perturbations.
... For instance, z plays a key role in antioxidant defense systems, DNA repair, and enzyme functio However, Cd disrupts Zn homeostasis in astrocytes by competing with Zn for sites, thereby displacing Zn and perturbing its normal functions [204,205]. This eve weakens the antioxidant defense of glial cells and causes CNS disease susceptibilit Also, considering that Zn is responsible for the functioning of metallothioneins (a that catalyzes the detoxification of metals against oxidative stress), the Cd-indu homeostasis imbalance can further lead to uncontrolled cellular damage that is imp in many neurodegenerative diseases [207,208]. ...
... This eventually weakens the antioxidant defense of glial cells and causes CNS disease susceptibility [206]. Also, considering that Zn is responsible for the functioning of metallothioneins (a protein that catalyzes the detoxification of metals against oxidative stress), the Cd-induced Zn homeostasis imbalance can further lead to uncontrolled cellular damage that is implicated in many neurodegenerative diseases [207,208]. ...
... Given the robust recapitulation of shMT2A sensitivity using encorafenib, it was of interest to evaluate the mechanistic connection to metallothioneins. Metallothioneins regulate reactive oxygen species (ROS) through regulation of zinc-finger transcription factors [32], mobilization of zinc [33], and contributing to the redox cycle of glutathione [34], the primary cytoplasmic antioxidant. ROS signals through the BRAF/CRAF-MEK-MAPK pathway, often upstream of RAF through RAS activation [35]. ...
... Scavenging ROS with NAC abolished the shMT2A selectivity of encorafenib and reduced cell loss ( Figure 7C). zinc-finger transcription factors [32], mobilization of zinc [33], and contributing to the redox cycle of glutathione [34], the primary cytoplasmic antioxidant. ROS signals through the BRAF/CRAF-MEK-MAPK pathway, often upstream of RAF through RAS activation [35]. ...
Background/Objectives: All 11 metallothionein protein-coding genes are located on human chromosome 16q13. It is unique among human genetics to have an entire pathway’s genes clustered in a short chromosomal region. Since solid tumors, particularly high-grade serous ovarian cancer (HGSC), exhibit high rates of monoallelic aneuploidy, this region is commonly lost. Studies have not yet been performed to determine what vulnerability may be created in cancer cells with low metallothionein expression. Here, a screen of FDA-approved cancer small molecule drugs for those best targeting low metallothionein ovarian cancer was completed. Methods: Screening methods were tested and compared using vehicle-treated negative controls and cadmium chloride, a positive control for cell loss selective for low metallothionein cells. CAOV3 cells, which are unique in their expression of only two metallothionein isoforms, were used, with or without shRNA knockdown of the predominantly expressed MT2A gene. A library of FDA-approved molecules was then screened. Results: The optimal assay utilized Hoechst 33342 nuclear staining and mechanized fluorescent microscope counting of cell content. Encorafenib, an RAF inhibitor, was identified as the most selective for enhanced cytotoxicity in MT2A knockdown cells compared to scrambled controls. Conclusions: The nuclear stain Hoechst 33342, assessed by fluorescence microscopy, provides a low variance, moderate throughput platform for cancer cell loss screens. Low metallothionein ovarian cancer cells exhibit a vulnerability to the RAF inhibitor encorafenib.
... These results are consistent with previous studies which showed that metallothionine proteins are involved in oxidative stress and inhibition of metallothionine proteins can increase cancer cell growth by regulating zinc-finger proteins. [25,26] To validate the RNA-seq results, we performed a subsequent RT-qPCR study in BxPC3 cells treated with DMSO, PK9328, or MS172 at different concentrations (Figure 5e). We observed a concentration-dependent upregulation of MT1X and MT1N genes and a simultaneous downregulation of ZNF391 and ZNF778 genes at 4 h time point induced by MS172, while PK9328 treatment led to no changes compared to the DMSO control. ...
Pharmacological reactivation of the tumor suppressor p53 remains a key challenge for the treatment of cancer. Acetylation Targeting Chimera (AceTAC), a novel technology is previously reported that hijacks lysine acetyltransferases p300/CBP to acetylate the p53Y220C mutant. However, p300/CBP are the only acetyltransferases harnessed for AceTAC development to date. In this study, it is demonstrated for the first time that the TAF1 acetyltransferase can be recruited to acetylate p53Y220C. A novel TAF1‐recruiting AceTAC, MS172 is discovered, which effectively acetylates p53Y220C lysine 382 in a concentration‐, time‐ and TAF1‐dependent manner via inducing the ternary complex formation between p53Y220C and TAF1. Notably, MS172 suppresses the proliferation in multiple p53Y220C‐harboring cancer cell lines more potently than the previously reported p300/CBP‐recruiting p53Y220C AceTAC MS78 with little toxicity in p53 WT and normal cells. Additionally, MS172 is bioavailable in mice and suitable for in vivo efficacy studies. Lastly, novel upregulation of metallothionine proteins by MS172‐induced p53Y220C acetylation is discovered using RNA‐seq and RT‐qPCR studies. This work demonstrates that TAF1 can be harnessed for AceTAC development and expands the very limited repertoire of the acetyltransferases that can be leveraged for developing AceTACs, thus advancing the targeted protein acetylation field.
