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Cys is one of the least abundant amino acids in proteins. However, it is often highly conserved and is usually found in important structural and functional regions of proteins. Its unique chemical properties allow it to undergo several post-translational modifications, many of which are mediated by reactive oxygen, nitrogen, sulfur, or carbonyl species. Thus, in addition to their role in catalysis, protein stability, and metal binding, Cys residues are crucial for the redox regulation of metabolism and signal transduction. In this review, we discuss Cys post-translational modifications (PTMs) and their role in plant metabolism and signal transduction. These modifications include the oxidation of the thiol group (S-sulfenylation, S-sulfinylation and S-sulfonylation), the formation of disulfide bridges, S-glutathionylation, persulfidation, S-cyanylation S-nitrosation, S-carbonylation, S-acylation, prenylation, CoAlation, and the formation of thiohemiacetal. For each of these PTMs, we discuss the origin of the modifier, the mechanisms involved in PTM, and their reversibility. Examples of the involvement of Cys PTMs in the modulation of protein structure, function, stability, and localization are presented to highlight their importance in the regulation of plant metabolic and signaling pathways. [ABSTRACT FROM AUTHOR]

Drug resistance is currently one of the biggest challenges in cancer treatment. With the deepening understanding of drug resistance, various mechanisms have been revealed, including metabolic reprogramming and alterations of redox balance. Notably, metabolic reprogramming mediates the survival of tumor cells in harsh environments, thereby promoting the development of drug resistance. In addition, the changes during metabolic pattern shift trigger reactive oxygen species (ROS) production, which in turn regulates cellular metabolism, DNA repair, cell death, and drug metabolism in direct or indirect ways to influence the sensitivity of tumors to therapies. Therefore, the intersection of metabolism and ROS profoundly affects tumor drug resistance, and clarifying the entangled mechanisms may be beneficial for developing drugs and treatment methods to thwart drug resistance. In this review, we will summarize the regulatory mechanism of redox and metabolism on tumor drug resistance and highlight recent therapeutic strategies targeting metabolic–redox circuits, including dietary interventions, novel chemosynthetic drugs, drug combination regimens, and novel drug delivery systems. [ABSTRACT FROM AUTHOR]

Drug resistance is currently one of the biggest challenges in cancer treatment. With the deepening understanding of drug resistance, various mechanisms have been revealed, including metabolic reprogramming and alterations of redox balance. Notably, metabolic reprogramming mediates the survival of tumor cells in harsh environments, thereby promoting the development of drug resistance. In addition, the changes during metabolic pattern shift trigger reactive oxygen species (ROS) production, which in turn regulates cellular metabolism, DNA repair, cell death, and drug metabolism in direct or indirect ways to influence the sensitivity of tumors to therapies. Therefore, the intersection of metabolism and ROS profoundly affects tumor drug resistance, and clarifying the entangled mechanisms may be beneficial for developing drugs and treatment methods to thwart drug resistance. In this review, we will summarize the regulatory mechanism of redox and metabolism on tumor drug resistance and highlight recent therapeutic strategies targeting metabolic–redox circuits, including dietary interventions, novel chemosynthetic drugs, drug combination regimens, and novel drug delivery systems.

Reactive oxygen species (ROS), products of normal cellular metabolism, play an important role in signal transduction. Autophagy is an intracellular degradation process in response to various stress conditions, such as nutritional deprivation, organelle damage and accumulation of abnormal proteins. ROS and autophagy both exhibit double-edged sword roles in the occurrence and development of cancer. Studies have shown that oxidative stress, as the converging point of these stimuli, is involved in the mechanical regulation of autophagy process. The regulation of ROS on autophagy can be roughly divided into indirect and direct methods. The indirect regulation of autophagy by ROS includes post-transcriptional and transcriptional modulation. ROS-mediated post-transcriptional regulation of autophagy includes the post-translational modifications and protein interactions of AMPK, Beclin 1, PI3K and other molecules, while transcriptional regulation mainly focuses on p62/Keap1/Nrf2 pathway. Notably, ROS can directly oxidize key autophagy proteins, such as ATG4 and p62, leading to the inhibition of autophagy pathway. In this review, we will elaborate the molecular mechanisms of redox regulation of autophagy in cancer, and discuss ROS- and autophagy-based therapeutic strategies for cancer treatment. [ABSTRACT FROM AUTHOR]

Cysteine protease ATG4B, a key autophagy protein, is an attractive target for colorectal cancer therapy. However, ATG4B inhibitors with higher efficiency, safety, and clear mechanism are still limited. In this study, we discovered ATG4B inhibitors based on the FDA-approved drug library through FRET-based high-throughput screening and gel-based analysis. Among the nine hits, compound Ebselen showed the most potent ATG4B inhibitory activity (IC50 = 189 nM) and exhibited controllable selectivity and structural optimizable possibility against ATG4A and caspases. We then performed mass spectrometry assay and cysteine mutations to confirm that Ebselen could covalently bind to ATG4B at Cys74. Moreover, Cys292 and Cys361 instead of Cys74 are responsible for the redox-oligomerization and efficient activity inhibition of ATG4B. Ultimately through cell culture and mouse xenograft tumor models, we established the impact of Ebselen on autophagy and tumor suppression via ATG4B inhibition other than apoptosis. These results suggest that old drug Ebselen as an ATG4B inhibitor through oxidative modification may be repurposed as a promising anti-colorectal cancer drug. Highlights: High-throughput screening of ATG4B inhibitors based on FDA-approved drug library Ebselen can covalently bind to ATG4B at Cys74 Ebselen can promote ATG4B oligomerization at Cys292 and Cys361 Ebselen suppresses the growth of colorectal cancer cells and xenograft tumors via ATG4B inhibition [ABSTRACT FROM AUTHOR]

Abstract Background S-glutathionylation is the formation of disulfide bonds between the tripeptide glutathione and cysteine residues of the protein, protecting them from irreversible oxidation and in some cases causing change in their functions. Regulatory glutathionylation of proteins is a controllable and reversible process associated with cell response to the changing redox status. Prediction of cysteine residues that undergo glutathionylation allows us to find new target proteins, which function can be altered in pathologies associated with impaired redox status. We set out to analyze this issue and create new tool for predicting S-glutathionylated cysteine residues. Results One hundred forty proteins with experimentally proven S-glutathionylated cysteine residues were found in the literature and the RedoxDB database. These proteins contain 1018 non-S-glutathionylated cysteines and 235 S-glutathionylated ones. Based on 235 S-glutathionylated cysteines, non-redundant positive dataset of 221 heptapeptide sequences of S-glutathionylated cysteines was made. Based on 221 heptapeptide sequences, a position-specific matrix was created by analyzing the protein sequence near the cysteine residue (three amino acid residues before and three after the cysteine). We propose the method for calculating the glutathionylation propensity score, which utilizes the position-specific matrix and a criterion for predicting glutathionylated peptides. Conclusion Non-S-glutathionylated sites were enriched by cysteines in − 3 and + 3 positions. The proposed prediction method demonstrates 76.6% of correct predictions of S-glutathionylated cysteines. This method can be used for detecting new glutathionylation sites, especially in proteins with an unknown structure.

Posttranslational modifications of protein cysteine thiols play a significant role in redox regulation and the pathogenesis of human diseases. Herein, we report the characterization of the cellular redox landscape in terms of quantitative, site-specific occupancies of both S-glutathionylation (SSG) and total reversible thiol oxidation (total oxidation) in RAW 264.7 macrophage cells under basal conditions. The occupancies of thiol modifications for ~4000 cysteine sites were quantified, revealing a mean site occupancy of 4.0% for SSG and 11.9% for total oxidation, respectively. Correlations between site occupancies and structural features such as pKa, relative residue surface accessibility, and hydrophobicity were observed. Proteome-wide site occupancy analysis revealed that the average occupancies of SSG and total oxidation in specific cellular compartments correlate well with the expected redox potentials of respective organelles in macrophages, consistent with the notion of redox compartmentalization. The lowest average occupancies were observed in more reducing organelles such as the mitochondria (non-membrane) and nucleus, while the highest average occupancies were found in more oxidizing organelles such as endoplasmic reticulum (ER) and lysosome. Furthermore, a pattern of subcellular susceptibility to redox changes was observed under oxidative stress induced by exposure to engineered metal oxide nanoparticles. Peroxisome, ER, and mitochondria (membrane) are the organelles which exhibit the most significant redox changes; while mitochondria (non-membrane) and Golgi were observed as the organelles being most resistant to oxidative stress. Finally, it was observed that Cys residues at enzymatic active sites generally had a higher level of occupancy compared to non-active Cys residues within the same proteins, suggesting site occupancy as a potential indicator of protein functional sites. The raw data are available via ProteomeXchange with identifier PXD019913.

Background: One of the fascinating aspects of epigenetic regulation is that it provides means to rapidly adapt to environmental change. This is particularly relevant in the plant kingdom, where most species are sessile and exposed to increasing habitat fluctuations due to global warming. Although the inheritance of epigenetically controlled traits acquired through environmental impact is a matter of debate, it is well documented that environmental cues lead to epigenetic changes, including chromatin modifications, that affect cell differentiation or are associated with plant acclimation and defense priming. Still, in most cases, the mechanisms involved are poorly understood. An emerging topic that promises to reveal new insights is the interaction between epigenetics and metabolism. Scope of review: This study reviews the links between metabolism and chromatin modification, in particular histone acetylation, histone methylation, and DNA methylation, in plants and compares them to examples from the mammalian field, where the relationship to human diseases has already generated a larger body of literature. This study particularly focuses on the role of reactive oxygen species (ROS) and nitric oxide (NO) in modulating metabolic pathways and gene activities that are involved in these chromatin modifications. As ROS and NO are hallmarks of stress responses, we predict that they are also pivotal in mediating chromatin dynamics during environmental responses. Major conclusions: Due to conservation of chromatin-modifying mechanisms, mammals and plants share a common dependence on metabolic intermediates that serve as cofactors for chromatin modifications. In addition, plant-specific non-CG methylation pathways are particularly sensitive to changes in folate-mediated one-carbon metabolism. Finally, reactive oxygen and nitrogen species may fine-tune epigenetic processes and include similar signaling mechanisms involved in environmental stress responses in plants as well as animals.

Redox modification of functional or regulatory proteins has emerged as an important mechanism of post-translational modification. However, the role of redox modifications of transcription factors mediated by methionine sulfoxide reductase (Msr) in regulating physiological processes in plants remains unclear, especially in fruit ripening. In this study, we determined that MaNAC42, a transcriptional activator, is involved in the regulation of fruit ripening in banana under oxidative stress. Integrated analysis of ChIP-qPCR and EMSA data showed that MaNAC42 directly binds to promoters of genes related to oxidative stress and ripening. Ectopic overexpression of MaNAC42 in Arabidopsis delays dark-induced senescence in leaves, indicating that MaNAC42 plays a negative role in senescence. Furthermore, we found that MaNAC42 is a target of MaMsrB2, a methionine sulfoxide reductase B. Methionine oxidation in MaNAC42 (i.e. sulfoxidation) or mimicking sulfoxidation by mutating methionine to glutamine both lead to decreased DNA-binding capacity and transcriptional activity. On the other hand, MaMsrB2 can partially repair oxidized MaNAC42 and restore its DNA-binding capacity. Thus, our results suggest a novel regulatory mechanism of fruit ripening in banana involving MaMsrB2-mediated redox regulation of the ripening-related transcription factor MaNAC42. [ABSTRACT FROM AUTHOR]

Either NPR1 or TGA1 serve as master redox-sensitive transcriptional regulators for the transcription of PR genes in plants. The redox modification of the two co-activators involved in BABA-induced priming resistance against Botrytis cinerea in grapes was examined in this study. The results showed that 10 mmol L−1 BABA could effectively trigger a priming defense in grapes as manifested by augmented expression levels of PR genes upon inoculation with B. cinerea. Moreover, transcriptome profiling analysis revealed that all of the sets of key genes in the enzymatic ROS scavenging system, the PPP and AsA-GSH cycle were in harmony and were transcriptionally induced in BABA-primed grapes with pathogenic infection; in addition, this enhanced expression caused the accelerated accumulation of reductive substances, namely, AsA, GSH and NADPH, resulting in reduced intercellular conditions. Under reduced conditions, the interaction of VvTGA1 and VvNPR1 in the Y2H assay implied that VvTGA1 can provide the DNA binding capacity required by VvNPR1 for activation of VvPR genes. Consequently, the transactivation of VvNPR1 by the promoters of VvPR1 , VvPR2 and VvPR5 was determined via a DLR assay, and it induced the transcription of the VvPR genes. In parallel, the redox-modified reducing condition achieved with an abundant supply of reductive substances was closely associated with the translocation of NPR1 for interaction with TGA in the nucleus. Thus, the posttranslational modification and subsequent interaction of the two redox-sensitive co-activators of VvNPR1 and VvTGA1 under reduced conditions may be responsible for BABA-induced priming for effective disease resistance in grapes. Image 1 • BABA-triggered resistance against B. cinerea in grapes contributed to a priming defence. • Maintenance of redox homeostasis by BABA is a backup mechanism for priming defense. • Interaction between NPR1 and TGA1 was essential for defence activation. • Assembling NPR1-TGA1 regulator depended on intercellular reduced status. [ABSTRACT FROM AUTHOR]

Diabetes mellitus is one of the major public health problems worldwide. Considerable recent evidence suggests that the cellular reduction-oxidation (redox) imbalance leads to oxidative stress and subsequent occurrence and development of diabetes and related complications by regulating certain signaling pathways involved in β-cell dysfunction and insulin resistance. Reactive oxide species (ROS) can also directly oxidize certain proteins (defined as redox modification) involved in the diabetes process. There are a number of potential problems in the clinical application of antioxidant therapies including poor solubility, storage instability and nonselectivity of antioxidants. Novel antioxidant delivery systems may overcome pharmacokinetic and stability problem and improve the selectivity of scavenging ROS. We have therefore focused on the role of oxidative stress and antioxidative therapies in the pathogenesis of diabetes mellitus. Precise therapeutic interventions against ROS and downstream targets are now possible and provide important new insights into the treatment of diabetes. [ABSTRACT FROM AUTHOR]

An electrochemical lateral flow immunoassay (eLFIA) strip with high reproducibility was developed to rapidly and accurately detect Streptococcus suis serotype 2. This proposed strip was fabricated by integrating ratiometric electrochemical detection and LFIA (R-eLFIA). The R-eLFIA exhibited excellent reproducibility, which was improved by 3.8 times compared to a single electrode. A dual-working screen-printed graphene electrode (SPGE) was designed by tuning the working electrode with electroactive species in the biosensing system. Ferrocene carboxylic acid (Fc) was used as a signal probe, and sunset yellow (SY) at one working electrode was used as an internal reference signal to provide a built-in correction for reducing the effects of inherent background current. S. suis serotype 2-specific antibodies were immobilized on a nitrocellulose membrane of LFIA, which is located on the position of Fc-SPGE. In the presence of the analyte, an immunocomplex formed on the region of Fc-SPGE, causing a decrease in Fc current while SY current remained constant. The current ratio's decrease was proportional to S. suis serotype 2's concentration. Under optimization, this biosensor showed good linearity in the range of 102–1010 CFU/mL with a limit of detection of 10 CFU/mL and achieved a rapid detection time (15 min). Moreover, the R-eLFIA biosensor exhibited excellent reproducibility and high selectivity and was applied in human serum samples. Thus, this study successfully matched the advantages of the ratiometric strategy and LFIA and has great potential to be used as an effective tool for rapidly detecting S. suis serotype 2 in clinical samples. [Display omitted] [ABSTRACT FROM AUTHOR]

We describe herein a synthetic strategy for the synthesis of new fluorescent imidazole and phenazine derivatives synthesized from lapachol, a naturally occurring naphthoquinone isolated from Tabebuia species (ipê tree). The photophysical properties and computational details of these compounds have been studied, aiming at gaining a complete understanding of the potential of these derivatives as probes capable of staining mitochondria selectively. Cell imaging experiments proved the capacity of the imidazole derivatives as selective fluorescent mitochondrial imaging probes. These heterocycles present the same staining patterns as MitoTracker Red, corroborating the potential of these compounds as new mitochondria markers permeable to the cell membrane. [ABSTRACT FROM AUTHOR]

The heart is able to respond acutely to changes in oxygen tension. Since ion channels can respond rapidly to stimuli, the “ion channel oxygen sensing hypothesis” has been proposed to explain acute adaptation of cells to changes in oxygen demand. However the exact mechanism for oxygen sensing continues to be debated. Mitochondria consume the lion’s share of oxygen in the heart, fuelling the production of ATP that drives excitation and contraction. Mitochondria also produce reactive oxygen species that are capable of altering the redox state of proteins. The cardiac L-type calcium channel is responsible for maintaining excitation and contraction. Recently, the reactive cysteine on the cardiac L-type calcium channel was identified. These data clarified that the channel does not respond directly to changes in oxygen tension, but rather responds to cellular redox state. This leads to acute alterations in cell signalling responsible for the development of arrhythmias and pathology. [ABSTRACT FROM AUTHOR]

Background Reactive oxygen species (ROS), the natural byproducts of aerobic metabolism, are precisely orchestrated to evoke diverse signaling pathways. To date, studies have focused mainly on the detrimental effects of ROS in stem cells. Recently, accumulating evidence has suggested that ROS also function as second messengers that modulate stem cell self-renewal and differentiation by regulating intricate signaling networks. Although many efforts have been made to clarify the general effects of ROS on signal transduction in stem cells, less is known about the initial and direct executors of ROS signaling, which are known as ‘redox sensors’. Scope of review Modifications of cysteine residues in redox sensors are of significant importance in the modulation of protein function in response to different redox conditions. Intriguingly, most key molecules in ROS signaling and cell cycle regulation (including transcriptional factors and kinases) that are crucial in the regulation of stem cell self-renewal and differentiation have the potential to be redox sensors. Major conclusions We highlight herein the importance of redox regulation of these key regulators in stem cell self-renewal and differentiation. General significance Understanding the mechanisms of redox regulation in stem cell self-renewal and differentiation will open exciting new perspectives for stem cell biology. This article is part of a Special Issue entitled Redox regulation of differentiation and de-differentiation. [ABSTRACT FROM AUTHOR]

High mobility group box 1 (HMGB1) is a nuclear protein with extracellular inflammatory cytokine activity. It is released passively during cell injury and necrosis, and secreted actively by immune cells. HMGB1 contains three conserved redox-sensitive cysteine residues: C23 and C45 can form an intramolecular disulfide bond, whereas C106 is unpaired and is essential for the interaction with Toll-Like Receptor (TLR) 4. However, a comprehensive characterization of the dynamic redox states of each cysteine residue and of their impacts on innate immune responses is lacking. Using tandem mass spectrometric analysis, we now have established that the C106 thiol and the C23-C45 disulfide bond are required for HMGB1 to induce nuclear NF-κB translocation and tumor necrosis factor (TNF) production in macrophages. Both irreversible oxidation to sulphonates and complete reduction to thiols of these cysteines inhibited TNF production markedly. In a proof of concept murine model of hepatic necrosis induced by acetaminophen, during inflammation, the predominant form of serum HMGB1 is the active one, containing a C106 thiol group and a disulfide bond between C23 and C45, whereas the inactive form of HMGB1, containing terminally oxidized cysteines, accumulates during inflammation resolution and hepatic regeneration. These results reveal critical posttranslational redox mechanisms that control the proinflammatory activity of HMGB1 and its inactivation during pathogenesis. [ABSTRACT FROM AUTHOR]

Aims: Although cardiac alternans is a known predictor of lethal arrhythmias, its underlying causes remain largely undefined in disease settings. The potential role of, and mechanisms responsible for, beat-to-beat alternations in the amplitude of systolic Ca2+ transients (Ca2+ alternans) was investigated in a canine post-myocardial infarction (MI) model of sudden cardiac death (SCD). [ABSTRACT FROM PUBLISHER]

Abstract: The tumor suppressor protein p53 is a redox-active transcription factor that organizes and directs cellular responses in the face of a variety of stresses that lead to genomic instability. One of the most important questions in the study of p53 is how selective transactivation of certain p53 target genes is achieved. Reactive oxygen species (ROS), generated by cells as products or by-products, can function either as signaling molecules or as cellular toxicants. Cellular generation of ROS is central to redox signaling. Recent studies have revealed that each cellular concentration and distribution of p53 has a distinct cellular function and that ROS act as both an upstream signal that triggers p53 activation and a downstream factor that mediates apoptosis. Here, we examine the newly discovered role of p53 in regulating cellular ROS generation and how ROS modulate selective transactivation of p53 target genes. The focus is on interlinks between ROS and p53. [Copyright &y& Elsevier]

Abstract: Nitric oxide (NO) modulates the release of various neurotransmitters, some of these are considered to be involved in neuronal plasticity that includes long-term depression in the cerebellum. To date, there have been no reports on the modulation of the exocytotic release of neurotransmitters in the cerebellar granule cells (CGCs) by NO. The aim of this study was to investigate the effects of NO on the exocytotic release of glutamate from rat CGCs. Treatment with NO-related reagents revealed that NO inhibited high-K+-evoked glutamate release. Clostridium botulinum type B neurotoxin (BoNT/B) attenuated the enhancement of glutamate release caused by NO synthase (NOS) inhibition; this indicates that NO acts on the high-K+-evoked exocytotic pathway. cGMP-related reagents did not affect the high-K+-evoked glutamate release. NO-related reagents did not affect Ca2+ ionophore-induced glutamate release, suggesting that NO inhibits Ca2+ entry through voltage-dependent Ca2+ channels (VDCC). Monitoring of intracellular Ca2+ revealed that NO inhibited high-K+-evoked Ca2+ entry. L-type VDCC blockers inhibited glutamate release and NO did not have an additive effect on the inhibition produced by the L-type VDCC blocker. The inhibition of the high-K+-evoked glutamate release by NO was abolished by a reducing reagent; this suggested that NO regulates the high-K+-evoked glutamate release from CGCs by redox modulation. [Copyright &y& Elsevier]

We examined hypoxia-induced changes in global thiol proteome profile in human prostate cancer cells using a BIAM-based display method. We analyzed the kinetics of protein thiol modification by using a pattern recognition algorithm, self-organizing maps (SOM) clustering, and identified the BIAM-labeled proteins by MALDI-TOF and ESI-tandem mass spectrometry. We found 99 out of 215 of total BIAM-labeled proteins were affected by hypoxia treatment and, yet, with diverse patterns and kinetics of redox modification. Our study proved that proteomics analysis employing the BIAM-labeling method can provide valuable information pertaining to global changes in the redox status of proteins in response to hypoxia. [ABSTRACT FROM AUTHOR]

Generation of a monomethylated selenium metabolite is critical for the anticancer activity of selenium. Because of its strong nucleophilicity, the metabolite can react directly with protein thiols to cause redox modification. Here, we report a neural network-based analysis to identify potential selenium targets. A reactive thiol specific reagent, BIAM, was used to monitor thiol proteome changes on 2D gel. We constructed a dynamic model and evaluated the relative importance of proteins mediating the cellular responses to selenium. Information from this study will provide new clues to unravel mechanisms of anticancer action of selenium. High impact selenium targets could also serve as biomarkers to gauge the efficacy of selenium chemoprevention. [ABSTRACT FROM AUTHOR]

Redox modification of proteins is proposed to play a central role in regulating cellular function. However, high-throughput techniques for the analysis of the redox status of individual proteins in complex mixtures are lacking. The aim was thus to develop a suitable technique to rapidly identify proteins undergoing oxidation of critical thiols by S-glutathionylation. The method is based on the specific reduction of mixed disulfides by glutaredoxin, their reaction with N-ethylmaleimide–biotin, affinity purification of tagged proteins, and identification by proteomic analysis. The method unequivocally identified 43 mostly novel cellular protein substrates for S-glutathionylation. These include protein chaperones, cytoskeletal proteins, cell cycle regulators, and enzymes of intermediate metabolism. Comparisons of the patterns of S-glutathionylated proteins extracted from cells undergoing diamide-induced oxidative stress and during constitutive metabolism reveal both common protein substrates and substrates failing to undergo enhanced S-glutathionylation during oxidative stress. The ability to chemically tag, select, and identify S-glutathionylated proteins, particularly during constitutive metabolism, will greatly enhance efforts to establish posttranslational redox modification of cellular proteins as an important biochemical control mechanism in coordinating cellular function. [Copyright &y& Elsevier]

Cancer cells utilize rapidly elevated cellular antioxidant programs to accommodate chemotherapy-induced oxidative stress; however, the underlying mechanism remains largely unexplored. Here we screen redox-sensitive effectors as potential therapeutic targets for colorectal cancer (CRC) treatment and find that cyclophilin A (CypA) is a compelling candidate. Our results show that CypA forms an intramolecular disulfide bond between Cys115 and Cys161 upon oxidative stress and the oxidized cysteines in CypA are recycled to a reduced state by peroxiredoxin-2 (PRDX2). Furthermore, CypA reduces cellular reactive oxygen species levels and increases CRC cell survival under insults of H 2 O 2 and chemotherapeutics through a CypA-PRDX2-mediated antioxidant apparatus. Notably, CypA is upregulated in chemoresistant CRC samples, which predicts poor prognosis. Moreover, targeting CypA by cyclosporine A exhibits promising efficacy against chemoresistant CRC when combined with chemotherapeutics. Collectively, our findings highlight CypA as a component of cellular noncanonical antioxidant defense and as a potential druggable therapeutic target to ameliorate CRC chemoresistance. [Display omitted] • CypA is identified as a redox-sensitive and druggable protein upregulated in CRC • CypA forms a disulfide bond at Cys115 and Cys161, which is reduced by PRDX2 • CypA elicits CRC chemoresistance via redox modification-mediated antioxidant defense • Targeting CypA by cyclosporine A potentiates chemotherapeutic efficacy in CRC Peng et al. show that CypA is a redox-sensitive effector regulating cellular oxidative status through a PRDX2-mediated noncanonical antioxidant system, which contributes to colorectal cancer (CRC) resistance to 5-fluorouracil and oxaliplatin. Targeting CypA by cyclosporine A sensitizes CRC to chemotherapy, indicating that CypA may serve as a potential druggable therapeutic target. [ABSTRACT FROM AUTHOR]

S-glutathionylation, the post-translational modification forming mixed disulfides between protein reactive thiols and glutathione, regulates redox-based signaling events in the cell and serves as a protective mechanism against oxidative damage. S-glutathionylation alters protein function, interactions, and localization across physiological processes, and its aberrant function is implicated in various human diseases. In this review, we discuss the current understanding of the molecular mechanisms of S-glutathionylation and describe the changing levels of expression of S-glutathionylation in the context of aging, cancer, cardiovascular, and liver diseases. [ABSTRACT FROM AUTHOR]

Background: S-glutathionylation is the formation of disulfide bonds between the tripeptide glutathione and cysteine residues of the protein, protecting them from irreversible oxidation and in some cases causing change in their functions. Regulatory glutathionylation of proteins is a controllable and reversible process associated with cell response to the changing redox status. Prediction of cysteine residues that undergo glutathionylation allows us to find new target proteins, which function can be altered in pathologies associated with impaired redox status. We set out to analyze this issue and create new tool for predicting S-glutathionylated cysteine residues. Results: One hundred forty proteins with experimentally proven S-glutathionylated cysteine residues were found in the literature and the RedoxDB database. These proteins contain 1018 non-S-glutathionylated cysteines and 235 S-glutathionylated ones. Based on 235 S-glutathionylated cysteines, non-redundant positive dataset of 221 heptapeptide sequences of S-glutathionylated cysteines was made. Based on 221 heptapeptide sequences, a position-specific matrix was created by analyzing the protein sequence near the cysteine residue (three amino acid residues before and three after the cysteine). We propose the method for calculating the glutathionylation propensity score, which utilizes the position-specific matrix and a criterion for predicting glutathionylated peptides. Conclusion: Non-S-glutathionylated sites were enriched by cysteines in − 3 and + 3 positions. The proposed prediction method demonstrates 76.6% of correct predictions of S-glutathionylated cysteines. This method can be used for detecting new glutathionylation sites, especially in proteins with an unknown structure. [ABSTRACT FROM AUTHOR]

One of the fascinating aspects of epigenetic regulation is that it provides means to rapidly adapt to environmental change. This is particularly relevant in the plant kingdom, where most species are sessile and exposed to increasing habitat fluctuations due to global warming. Although the inheritance of epigenetically controlled traits acquired through environmental impact is a matter of debate, it is well documented that environmental cues lead to epigenetic changes, including chromatin modifications, that affect cell differentiation or are associated with plant acclimation and defense priming. Still, in most cases, the mechanisms involved are poorly understood. An emerging topic that promises to reveal new insights is the interaction between epigenetics and metabolism. This study reviews the links between metabolism and chromatin modification, in particular histone acetylation, histone methylation, and DNA methylation, in plants and compares them to examples from the mammalian field, where the relationship to human diseases has already generated a larger body of literature. This study particularly focuses on the role of reactive oxygen species (ROS) and nitric oxide (NO) in modulating metabolic pathways and gene activities that are involved in these chromatin modifications. As ROS and NO are hallmarks of stress responses, we predict that they are also pivotal in mediating chromatin dynamics during environmental responses. Due to conservation of chromatin-modifying mechanisms, mammals and plants share a common dependence on metabolic intermediates that serve as cofactors for chromatin modifications. In addition, plant-specific non-CG methylation pathways are particularly sensitive to changes in folate-mediated one-carbon metabolism. Finally, reactive oxygen and nitrogen species may fine-tune epigenetic processes and include similar signaling mechanisms involved in environmental stress responses in plants as well as animals. • Plant specific non-CG methylation pathways are particularly sensitive to changes in folate-mediated one-carbon metabolism. • ROS and NO coordinate metabolic pathways and gene activities involved in chromatin modulation. • ROS and NO might also be pivotal in mediating chromatin dynamics during environmental stress responses. • Mammals and plants share a common dependence on metabolic intermediates that serve as cofactors for chromatin modulation. [ABSTRACT FROM AUTHOR]

Two Cys residues located in the α5-subunit of the yeast 20 S proteasome (20SPT) are post-translationally modified by S-glutathionylation. Such modification implies on the opening of the 20SPT gate when compared to the DTT-reduced purified preparations. The site-specific mutation of the highly conserved (yeast to human) α5-Cys76 residue induced either decreased cell viability or life span when compared to the wild type strain. The α5-C76S-mutated strain presented decreased resistance to oxidative stress without any alteration on major redox parameters. The closed gate conformation of the 20SPT pool in that short-lived strain represents 70% of total 20SPT pool against 35% in the wild type counterpart. Mass spectrometry analysis of the α5-subunit in the α5-C76S-mutated strain showed S-glutathionylation of the remaining α5-C221 residue suggesting that redox modification of the conserved α5-C76, but not of the α5-C221, is the one involved in the gate opening. Circular dichroism analyses of recombinant α5-WT and α5-C76S subunits showed similar pattern between them regarding their secondary and tertiary structures. The differences between the wild type and short-lived strains regarding prevalence of 20SPT gate conformation, clearly demonstrate that the 20SPT opening is relevant to maintain cellular homeostasis and life span. [ABSTRACT FROM AUTHOR]

Upregulations of neuronal nitric oxide synthase (nNOS) in the rodent brain have been associated with neuronal aging. To address underlying mechanisms we generated SH-SY5Y neuronal cells constitutively expressing nNOS at a level similar to mouse brain (nNOS+ versus MOCK). Initial experiments revealed S-nitrosylations (SNO) of key players of protein homeostasis: heat shock cognate HSC70/HSPA8 within its nucleotide-binding site, and UBE2D ubiquitin conjugating enzymes at the catalytic site cysteine. HSPA8 is involved in protein folding, organelle import/export and chaperone-mediated LAMP2a-dependent autophagy (CMA). A set of deep redox and full proteome analyses, plus analysis of autophagy, CMA and ubiquitination with rapamycin and starvation as stimuli confirmed the initial observations and revealed a substantial increase of SNO modifications in nNOS+ cells, in particular targeting protein networks involved in protein catabolism, ubiquitination, carbohydrate metabolism and cell cycle control. Importantly, NO-independent reversible oxidations similarly occurred in both cell lines. Functionally, nNOS caused an accumulation of proteins, including CMA substrates and loss of LAMP2a. UBE2D activity and proteasome activity were impaired, resulting in dysregulations of cell cycle checkpoint proteins. The observed changes of protein degradation pathways caused an expansion of the cytoplasm, large lysosomes, slowing of the cell cycle and suppression of proliferation suggesting a switch of the phenotype towards aging, supported by downregulations of neuronal progenitor markers but increase of senescence-associated proteins. Hence, upregulation of nNOS in neuronal cells imposes aging by SNOing of key players of ubiquitination, chaperones and of substrate proteins leading to interference with crucial steps of protein homeostasis., (Copyright © 2018. Published by Elsevier B.V.)

Catechol estrogens (CEs) are metabolic electrophiles that actively undergo covalent interaction with cellular proteins, influencing molecular function. There is no feasible method to identify their binders in a living system. Herein, we developed a click chemistry-based approach using ethinylestradiol (EE2) as the precursor probe coupled with quantitative proteomics to identify protein targets of CEs and classify their binding strengths. Using in situ metabolic conversion and click reaction in liver microsomes, CEs-protein complex was captured by the probe, digested by trypsin, stable isotope labeled via reductive amination, and analyzed by liquid chromatography-mass spectrometry (LC-MS). A total of 334 liver proteins were repeatedly identified ( n ≥ 2); 274 identified proteins were classified as strong binders based on precursor mass mapping. The binding strength was further scaled by D/H ratio (activity probe/solvent): 259 strong binders had D/H > 5.25; 46 weak binders had 5.25 > D/H > 1; 5 nonspecific binders (keratins) had D/H < 1. These results were confirmed using spiked covalent control (strong binder) and noncovalent control (weak binder), as well as in vitro testing of cytochrome c (D/H = 5.9), which showed covalent conjugation with CEs. Many identified strong binders, such as glutathione transferase, catechol-O-methyl transferase, superoxide dismutase, catalase, glutathione peroxidase, and cytochrome c, are involved in cellular redox processes or detoxification activities. CE conjugation was shown to suppress the superoxide oxidase activity of cytochrome c, suggesting that CEs modification may alter the redox action of cellular proteins. Due to structural similarity and inert alkyne group, EE2 probe is very likely to capture protein targets of CEs in general. Thus, this strategy can be adopted to explore the biological impact of CEs modification in living systems.

High mobility group box-1 (HMGB1) is a nuclear protein that is expressed in almost all eukaryotic cells. In the nucleus, it maintains nuclear homeostasis and promotes gene transcription. HMGB1 can be passively released into the extracellular milieu after cell necrosis or actively secreted by activated immune cells. HMGB1 has several receptors such as Toll-like receptor 2, Toll-like receptor 4, and the receptor for advanced glycation end products. After brain injury, HMGB1 is released early from neural cells and contributes to the initial stages of the inflammatory response. However, surprisingly, HMGB1 can mediate beneficial effects during the course of stroke recovery. The biphasic biological property of extracellular HMGB1 may be related to the redox modifications of its cysteine residues. This review discusses the emerging roles of HMGB1 in several stroke models, as well as its potential role as a therapeutic target for stroke patients., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.)

Background: The generation of a monomethylated selenium metabolite is critical for the anticancer activity of selenium. Because of its strong nucleophilicity, the metabolite can react directly with protein thiols to cause redox modification. These chemical changes have never been examined systematically before because of the lack of a reliable methodology to study reactive protein thiols globally in cells and to quantify their redox status., Materials and Methods: PC-3 human prostate cancer cells were treated with methylseleninic acid (MSA) for 0.5, 1, 2, 3, 6, 12 or 24 h. A reactive thiol specific reagent, BIAM, was used to detect the extent of global redox changes on a 2D gel electrophoresis display. The data were analyzed by the Self Organizing Maps clustering algorithm. Protein identification was done by MALDI-TOF and ESI-tandem mass spectrometry., Results: Out of a total of 194 reactive thiol-containing protein spots on the 2D gel display, 100 of them (cluster 1) were not sensitive to MSA modulation. The remaining 94 were categorized into three distinct patterns. Cluster 2 (60 proteins) showed an immediate and sustained loss of reactive thiols for at least 24 h; cluster 3 (19 proteins) showed a transient loss of reactive thiols followed by a rapid rebound; and cluster 4 (15 proteins) showed a transient gain followed by a rapid return to normal. In contrast, there were minimal protein redox changes in control cells (not treated with MSA) over the same period of time. A total of 85 proteins were identified of which 40 were in clusters 2 to 4. The proteins which are sensitive to redox modification by MSA are distributed in various subcellular compartments. Western blot analysis showed that a number of chaperones were significantly induced by MSA., Conclusion: Global redox modification of proteins can be a major driving force of cellular stress, since these changes are likely to lead to protein unfolding, misfolding or aggregation. The induction of chaperones in cells treated with MSA is consistent with this interpretation since chaperones are charged with rescuing misfolded proteins. The above scenario is discussed in relation to an adaptive response which ultimately determines how cells respond to treatment with selenium., (Copyright© 2005 International Institute of Anticaner Research (Dr. John G. Delinassios), All rights reserved.)