Glutathione and Glutaredoxin—Key Players in Cellular Redox Homeostasis and Signaling

This Special Issue of Antioxidants on Glutathione (GSH) and Glutaredoxin (Grx) was designed to collect review articles and original research studies focused on advancing the current understanding of the roles of the GSH/Grx system in cellular homeostasis and disease processes. The tripeptide glutathione (GSH) is the most abundant non-enzymatic antioxidant/nucleophilic molecule in cells. In addition to various metabolic reactions involving GSH and its oxidized counterpart GSSG, oxidative post-translational modification (PTM) of proteins has been a focal point of keen interest in the redox field over the last few decades. In particular, the S-glutathionylation of proteins (protein-SSG formation), i.e., mixed disulfides between GSH and protein thiols, has been studied extensively. This reversible PTM can act as a regulatory switch to interconvert inactive and active forms of proteins, thereby mediating cell signaling and redox homeostasis. The unique architecture of the GSH molecule enhances its relative abundance in cells and contributes to the glutathionyl specificity of the primary catalytic activity of the glutaredoxin enzymes, which play central roles in redox homeostasis and signaling, and in iron metabolism in eukaryotes and prokaryotes under physiological and pathophysiological conditions. The class-1 glutaredoxins are characterized as cytosolic GSH-dependent oxidoreductases that catalyze reversible protein S-glutathionylation specifically, thereby contributing to the regulation of redox signal transduction and/or the protection of protein thiols from irreversible oxidation. This Special Issue includes nine other articles: three original studies and six review papers. Together, these ten articles support the central theme that GSH/Grx is a unique system for regulating thiol-redox hemostasis and redox-signal transduction, and the dysregulation of the GSH/Grx system is implicated in the onset and progression of various diseases involving oxidative stress. Within this context, it is important to appreciate the complementary functions of the GSH/Grx and thioredoxin systems not only in thiol-disulfide regulation but also in reversible S-nitrosylation. Several potential clinical applications have emerged from a thorough understanding of the GSH/Grx redox regulatory system at the molecular level, and in various cell types in vitro and in vivo, including, among others, the concept that elevating Grx content/activity could serve as an anti-fibrotic intervention; and discovering small molecules that mimic the inhibitory effects of S-glutathionylation on dimer association could identify novel anti-viral agents that impact the key protease activities of the HIV and SARS-CoV-2 viruses. Thus, this Special Issue on Glutathione and Glutaredoxin has focused attention and advanced understanding of an important aspect of redox biology, as well as spawning questions worthy of future study

Regulation of DJ-1 by glutaredoxin 1 \u3ci\u3ein vivo – implications for Parkinson’s disease\u3c/i\u3e

Parkinson’s disease (PD) is the second most common neurodegenerative disease worldwide, caused by the degeneration of the dopaminergic neurons in the substantia nigra. Mutations in PARK7 (DJ-1) result in early onset autosomal recessive PD, and oxidative modification of DJ-1 has been reported to regulate the protective activity of DJ-1 in vitro. Glutathionylation is a prevalent redox modification of proteins resulting from the disulfide adduction of the glutathione moiety to a reactive cysteine-SH; and glutathionylation of specific proteins has been implicated in regulation of cell viability. Glutaredoxin 1 (Grx1) is the principal deglutathionylating enzyme within cells, and it has been reported to mediate protection of dopaminergic neurons in C. elegans, however many of the functional downstream targets of Grx1 in vivo remain unknown. Previously, DJ-1 protein content was shown to decrease concomitantly with diminution of Grx1 protein content in cell culture of model neurons (SH-SY5Y and Neuro-2A lines). In the current study we aimed to investigate the regulation of DJ-1 by Grx1 in vivo and characterize its glutathionylation in vitro. Here, with Grx−/− mice we provide evidence that Grx1 regulates protein levels of DJ-1 in vivo. Furthermore, with model neuronal cells (SH-SY5Y) we observed decreased DJ-1 protein content in response to treatment with known glutathionylating agents; and with isolated DJ-1 we identified two distinct sites of glutathionylation. Finally, we found that overexpression of DJ-1 in the dopaminergic neurons partly compensates for the loss of the Grx1 homolog in a C. elegans in vivo model of PD. Therefore; our results reveal a novel redox modification of DJ-1 and suggest a novel regulatory mechanism for DJ-1 content in vivo

HSV Infection Induces Production of ROS, which Potentiate Signaling from Pattern Recognition Receptors: Role for S-glutathionylation of TRAF3 and 6

The innate immune response constitutes the first line of defense against infections. Pattern recognition receptors recognize pathogen structures and trigger intracellular signaling pathways leading to cytokine and chemokine expression. Reactive oxygen species (ROS) are emerging as an important regulator of some of these pathways. ROS directly interact with signaling components or induce other post-translational modifications such as S-glutathionylation, thereby altering target function. Applying live microscopy, we have demonstrated that herpes simplex virus (HSV) infection induces early production of ROS that are required for the activation of NF-κB and IRF-3 pathways and the production of type I IFNs and ISGs. All the known receptors involved in the recognition of HSV were shown to be dependent on the cellular redox levels for successful signaling. In addition, we provide biochemical evidence suggesting S-glutathionylation of TRAF family proteins to be important. In particular, by performing mutational studies we show that S-glutathionylation of a conserved cysteine residue of TRAF3 and TRAF6 is important for ROS-dependent activation of innate immune pathways. In conclusion, these findings demonstrate that ROS are essential for effective activation of signaling pathways leading to a successful innate immune response against HSV infection

Glutathione and Glutaredoxin—Key Players in Cellular Redox Homeostasis and Signaling

This Special Issue of Antioxidants on Glutathione (GSH) and Glutaredoxin (Grx) was designed to collect review articles and original research studies focused on advancing the current understanding of the roles of the GSH/Grx system in cellular homeostasis and disease processes. The tripeptide glutathione (GSH) is the most abundant non-enzymatic antioxidant/nucleophilic molecule in cells. In addition to various metabolic reactions involving GSH and its oxidized counterpart GSSG, oxidative post-translational modification (PTM) of proteins has been a focal point of keen interest in the redox field over the last few decades. In particular, the S-glutathionylation of proteins (protein-SSG formation), i.e., mixed disulfides between GSH and protein thiols, has been studied extensively. This reversible PTM can act as a regulatory switch to interconvert inactive and active forms of proteins, thereby mediating cell signaling and redox homeostasis. The unique architecture of the GSH molecule enhances its relative abundance in cells and contributes to the glutathionyl specificity of the primary catalytic activity of the glutaredoxin enzymes, which play central roles in redox homeostasis and signaling, and in iron metabolism in eukaryotes and prokaryotes under physiological and pathophysiological conditions. The class-1 glutaredoxins are characterized as cytosolic GSH-dependent oxidoreductases that catalyze reversible protein S-glutathionylation specifically, thereby contributing to the regulation of redox signal transduction and/or the protection of protein thiols from irreversible oxidation. This Special Issue includes nine other articles: three original studies and six review papers. Together, these ten articles support the central theme that GSH/Grx is a unique system for regulating thiol-redox hemostasis and redox-signal transduction, and the dysregulation of the GSH/Grx system is implicated in the onset and progression of various diseases involving oxidative stress. Within this context, it is important to appreciate the complementary functions of the GSH/Grx and thioredoxin systems not only in thiol-disulfide regulation but also in reversible S-nitrosylation. Several potential clinical applications have emerged from a thorough understanding of the GSH/Grx redox regulatory system at the molecular level, and in various cell types in vitro and in vivo, including, among others, the concept that elevating Grx content/activity could serve as an anti-fibrotic intervention; and discovering small molecules that mimic the inhibitory effects of S-glutathionylation on dimer association could identify novel anti-viral agents that impact the key protease activities of the HIV and SARS-CoV-2 viruses. Thus, this Special Issue on Glutathione and Glutaredoxin has focused attention and advanced understanding of an important aspect of redox biology, as well as spawning questions worthy of future study

Dysregulation of Glutathione Homeostasis in Neurodegenerative Diseases

Dysregulation of glutathione homeostasis and alterations in glutathione-dependent enzyme activities are increasingly implicated in the induction and progression of neurodegenerative diseases, including Alzheimer’s, Parkinson’s and Huntington’s diseases, amyotrophic lateral sclerosis, and Friedreich’s ataxia. In this review background is provided on the steady-state synthesis, regulation, and transport of glutathione, with primary focus on the brain. A brief overview is presented on the distinct but vital roles of glutathione in cellular maintenance and survival, and on the functions of key glutathione-dependent enzymes. Major contributors to initiation and progression of neurodegenerative diseases are considered, including oxidative stress, protein misfolding, and protein aggregation. In each case examples of key regulatory mechanisms are identified that are sensitive to changes in glutathione redox status and/or in the activities of glutathione-dependent enzymes. Mechanisms of dysregulation of glutathione and/or glutathione-dependent enzymes are discussed that are implicated in pathogenesis of each neurodegenerative disease. Limitations in information or interpretation are identified, and possible avenues for further research are described with an aim to elucidating novel targets for therapeutic interventions. The pros and cons of administration of N-acetylcysteine or glutathione as therapeutic agents for neurodegenerative diseases, as well as the potential utility of serum glutathione as a biomarker, are critically evaluated

Targeted disruption of the glutaredoxin 1 gene does not sensitize adult mice to tissue injury induced by ischemia/reperfusion and hyperoxia

To understand the physiological function of glutaredoxin, a thiotransferase catalyzing the reduction of mixed disulfides of protein and glutathione, we generated a line of knockout mice deficient in the cytosolic glutaredoxin 1 (Grx1). To our surprise, mice deficient in Grx1 were not more susceptible to acute oxidative insults in models of heart and lung injury induced by ischemia/reperfusion and hyperoxia, respectively, suggesting that either changes in S-glutathionylation status of cytosolic proteins are not the major cause of such tissue injury or developmental adaptation in the Glrx1-knockout animals alters the response to oxidative insult. In contrast, mouse embryonic fibroblasts (MEFs) isolated from Grx1-deficient mice displayed an increased vulnerability to diquat and paraquat, but they were not more susceptible to cell death induced by hydrogen peroxide (H2O2) and diamide. A deficiency in Grx1 also sensitized MEFs to protein S-glutathionylation in response to H2O2 treatment and retarded deglutathionylation of the S-glutathionylated proteins, especially for a single prominent protein band. Additional experiments showed that MEFs lacking Grx1 were more tolerant to apoptosis induced by tumor necrosis factor αplus actinomycin D. These findings suggest that various oxidants may damage the cells via distinct mechanisms in which the action of Grx1 may or may not be protective and Grx1 may exert its function on specific target proteins