ReviewGlutathione synthesis and its role in redox signaling☆
Highlights
► Glutathione (GSH) is the most abundant antioxidant and a major detoxification agent in cells. ► Glutamate cysteine ligase, which catalyzes the first step in GSH synthetase is regulated by redox signaling. ► Accumulating evidence suggests that GSH plays important roles in cell signaling. ► Protein S-glutathionylation and S-nitrosylation play major roles in redox signaling.
Introduction
Several reactive species derived from oxygen and nitrogen are produced in cells from a variety sources such as NADPH oxidases (NOX) [1], leaks from the mitochondrial electron transport chain [2], redox cycling of quinones [3], and nitric oxide synthases (NOS) [4]. The principal reactive species produced enzymatically are superoxide (O2−), hydrogen peroxide (H2O2), and nitric oxide (NO). These can be involved in cell injury but are also the principal actors in cell signaling, particularly the latter two [5], [6], [7]. Other species derived from these, hydroxyl radical (OH) and peroxynitrous acid (ONOOH), are extremely potent oxidants and are more likely involved in cell injury than signaling. Environmental exposure adds additional reactive species such as ozone and nitrogen dioxide [8]. Together these species are often referred to as reactive oxygen (ROS) and nitrogen species [9], terms we will use sparingly as they are not helpful in understanding mechanism. Indeed, an argument has been made for considering only H2O2 as the actual species involved among the ROS while NO and nitrosoglutathione (GSNO), a conjugate of NO and glutathione (GSH) that is produced by an as yet not definitively demonstrated oxidative mechanism, are responsible for ROS and RNS signaling. Finally, other reactive species, lipid hydroperoxides (ROOH) and α,β-unsaturated aldehydes, that are derived from lipid peroxidation can also participate in both injury and cell signaling [7].
As mentioned, O2−, H2O2, and NO can contribute to pathology as well as signaling. Therefore, it is necessary for cells to maintain a narrow rate of production of these reactive species that is not directly harmful yet effective in mediating diverse physiologic functions through signaling.
Our focus here will be on the involvement of H2O2 and GSNO in signaling. The propensity of H2O2 to modify protein function is attributed to its ability to react with particular cysteine residues within proteins [10]. The possible oxidative modification of cysteine residues within proteins includes sulfenic acid (RSOH), intra- or intermolecular disulfide (RSSR), and S-glutathionylation (RSSG), sulfinic acid (RS(O)OH) and sulfonic acid (RS(O)2OH) acids, thiyl radicals (RS•), sulfenyl-amides, thiosulfinates, disulfide-S-monoxides; however, physiological signaling probably only involves the first three. Formation of the other species would influence signaling by producing species that are far more difficult to reduce back to a thiol than RSOH, RSSR and RSSG but are also less likely to form at low H2O2 concentration. Formation of RSOH, RSSG and RSSG can be transient and reversible, allowing these species to participate in biochemical functions, such as redox sensing and responding, catalysis, and signal transduction [11]. This redox-modification-based mode of signal transduction is called redox signaling. Only some select cysteine residues are involved in reactions with H2O2 or ROOH as even the more nucleophilic thiolate (S−) form is not strong enough to act in the absence of a nearby proton donor or metal to remove the OH− that would be the leaving group if a thiolate alone reacted with ROOH or H2O2. This complex chemistry has been recently reviewed [5].
The concentration of GSH, γ-l-glutamyl-l-cysteinyl-glycine, which is the most abundant non-protein thiol in cells, is in the range of 1–10 mM in most mammalian cells [12]. GSH is also the most abundant antioxidant and a major detoxification agent in cells. Enzymes such as glutathione peroxidases (GPx) and one of the peroxiredoxins (Prdx VI) catalyze the reduction of H2O2 (or ROOH) by GSH into H2O (or the corresponding alcohol (ROH) and GSSG). GSSG is reduced back to GSH by glutathione reductase using NADPH to maintain a steady state GSH/GSSG ratio that is almost all GSH but varies with cell type and disease state [13]. GSH also conjugates with electrophiles and thus participates in the metabolism and detoxification of endogenous compounds and xenobiotic toxicants [14], [15], [16]. In addition, GSH is involved in many other metabolic reactions [9], [17], [18]. Therefore, it is not surprising that GSH plays roles in various cellular processes such as cell growth, proliferation, and apoptosis. Thus, it is clear that GSH is also involved in cell signaling through which it affects these important cell functions.
GSH participates in cell signaling through at least two mechanisms, protein S-glutathionylation and cysteine S-nitrosylation through thiol exchange with GSNO. The former is formed when GSH conjugates with reactive cysteine residues within proteins to form protein mixed disulfides (PSSG), and the latter is formed by reaction of a thiolate (S−) with GSNO. Emerging evidence suggest that both are controlled redox signaling mechanisms. How GSNO is formed has been the subject of intense investigation and several mechanisms have been proposed [6], [19]. Of course, GSH may also indirectly participate in the redox signaling by changing cellular redox homeostasis. The rest of this review will focus on the participation of protein S-glutathionylation and GSNO in signaling and on how cells regulate GSH homeostasis.
Section snippets
Regulation of glutathione content
In most cells, GSH is synthesized de novo through a two-step reaction. First γ-glutamylcysteine is formed from glutamate and cysteine catalyzed by glutamate cysteine ligase (GCL). Then glycine is added by glutathione synthetase to form GSH. GCL activity and cysteine availability are two rate-limiting factors in GSH synthesis. It is apparently so important for cells to maintain redox homeostasis and normal cellular function that both its concentration and GSH/GSSG ratio are tightly regulated.
Regulation of GCL expression
The regulation of both subunits of GCL, the catalytic subunit (GCLC) and the modifier subunit (GCLM), has been extensively studied in the past 20 years. A variety of signaling pathways, such as ERK1/2 [28], JNK1/2 [29], p38MAPK [30], PKC [31], and PI3K [32], [33], and transcription factors, such as c-Jun [29], NF-κB [27], JunD [34], and Nrf2 [25], etc., have been found to be involved in the regulation of GCLC and GCLM genes. The most significant and intriguing finding though, is that both genes
Protein S-glutathionylation
In a recent review [57], several reaction mechanisms of protein S-glutathionylation (PSSG) formation were proposed. We have added to and modified these in accord with the chemistry of cysteine oxidation in signaling proteins that was discussed in our recent review [5] (Fig. 1). Proton catalyzed nucleophilic substitution: To break the O-O bond in ROOH bond (where R is either a H or a lipid) and form a protein-cysteine sulfenic acid and ROH, there must be two conditions met; (a) the cysteine must
GSNO: reaction and regulation
NO is involved in a wide range of biochemical reactions and cell signaling pathways through protein modification. Although NO can diffuse freely, it is readily oxidized and this limits its function as a second messenger. Formation of S-nitrosothiols (SNO) from NO and cysteine can protect NO from oxidative consumption and thereby extend NO bioavailability, both temporally and spatially. Protein S-nitrosylation can alter protein function and mediate signaling events and it has been considered as
Future direction
After decades of studies, it has been well established that redox-dependent reversible oxidative modification of proteins can occur under various pathophysiologic conditions, and is an important cell-signaling mode analogous to phosphorylation-based cell signaling mechanism. As the most abundant non-protein antioxidant in the cells, GSH is critical to maintain redox homeostasis and thus indirectly involved in redox signaling. Accumulating evidence suggest that GSH may directly participate in
References (115)
- et al.
The utility of superoxide dismutase in studying free radical reactions. II. The mechanism of the mediation of cytochrome c reduction by a variety of electron carriers
Journal of Biological Chemistry
(1970) - et al.
Chemical biology of nitric oxide: Insights into regulatory, cytotoxic, and cytoprotective mechanisms of nitric oxide
Free Radical Biology and Medicine
(1998) - et al.
The chemistry of cell signaling by reactive oxygen and nitrogen species and 4-hydroxynonenal
Archives of Biochemistry and Biophysics
(2008) - et al.
Human health effects of air pollution
Environmental Pollution
(2008) Glutathione metabolism and its selective modification
Journal of Biological Chemistry
(1988)- et al.
Glutathione adducts of oxyeicosanoids
Prostaglandins and Other Lipid Mediators
(2002) - et al.
Mechanisms of cystic fibrosis transmembrane conductance regulator activation by S-nitrosoglutathione
Journal of Biological Chemistry
(2006) - et al.
Catalytic and regulatory properties of the heavy subunit of rat kidney γ-glutamylcysteine synthetase
Journal of Biological Chemistry
(1993) - et al.
Mechanisms of glutamate cysteine ligase (GCL) induction by 4-hydroxynonenal
Free Radical Biology and Medicine
(2005) - et al.
NRF2 as a determinant of cellular resistance in retinoic acid cytotoxicity
Free Radical Biology and Medicine
(2008)
4-hydroxynonenal induces glutamate cysteine ligase through JNK in HBE1 cells
Free Radical Biology and Medicine
Inhibition of ERK and p38 MAP kinases inhibits binding of Nrf2 and induction of GCS genes
Biochemical and Biophysical Research Communications
Role of protein kinase C delta in curcumin-induced antioxidant response element-mediated gene expression in human monocytes
Biochemical and Biophysical Research Communications
Phosphorylation of Nrf2 at Ser40 by protein kinase C in response to antioxidants leads to the release of Nrf2 from INrf2: but is not required for Nrf2 stabilization/accumulation in the nucleus and transcriptional activation of antioxidant response element-mediated NAD(P)H:quinone oxidoreductase-1 gene expression
Journal of Biological Chemistry
Molecular mechanism activating Nrf2-Keap1 pathway in regulation of adaptive response to electrophiles
Free Radical Biology and Medicine
GSK-3beta acts upstream of Fyn kinase in regulation of nuclear export and degradation of NF-E2 related factor 2
Journal of Biological Chemistry
The role of c-Jun phosphorylation in EpRE activation of phase II genes
Free Radical Biology and Medicine
Nrf2 and c-Jun regulation of antioxidant response element (ARE)-mediated expression and induction of gamma-glutamylcysteine synthetase heavy subunit gene
Biochemical Pharmacology
Small maf (MafG and MafK) proteins negatively regulate antioxidant response element-mediated expression and antioxidant induction of the NAD(P)H:Quinone oxidoreductase1 gene
Journal of Biological Chemistry
TCF11/Nrf1 overexpression increases the intracellular glutathione level and can transactivate the gamma-glutamylcysteine synthetase (GCS) heavy subunit promoter
Biochimica et Biophysica Acta
Bach1 competes with Nrf2 leading to negative regulation of the antioxidant response element (ARE)-mediated NAD(P)H:quinone oxidoreductase 1 gene expression and induction in response to antioxidants
Journal of Biological Chemistry
Nrf3 negatively regulates antioxidant-response element-mediated expression and antioxidant induction of NAD(P)H:quinone oxidoreductase1 gene
Journal of Biological Chemistry
Identification of activating transcription factor 4 (ATF4) as an Nrf2-interacting protein. Implication for heme oxygenase-1 gene regulation
Journal of Biological Chemistry
Functional characterization of transcription regulators that interact with the electrophile response element
Biochemical and Biophysical Research Communications
Molecular mechanism of the regulation of glutathione synthesis by tumor necrosis factor-alpha and dexamethasone in human alveolar epithelial cells
Journal of Biological Chemistry
Glutathiolation of proteins by glutathione disulfide S-oxide derived from S-nitrosoglutathione. Modifications of rat brain neurogranin/RC3 and neuromodulin/GAP-43
Journal of Biological Chemistry
Glutathionylation of proteins by glutathione disulfide S-oxide
Biochemical Pharmacology
Novel role for glutathione S-transferase pi. Regulator of protein S-Glutathionylation following oxidative and nitrosative stress
Journal of Biological Chemistry
Targeted disruption of the glutaredoxin 1 gene does not sensitize adult mice to tissue injury induced by ischemia/reperfusion and hyperoxia
Free Radical Biology and Medicine
Glutaredoxin 2 knockout increases sensitivity to oxidative stress in mouse lens epithelial cells
Free Radical Biology and Medicine
Glutathione supplementation potentiates hypoxic apoptosis by S-glutathionylation of p65-NFkappaB
Journal of Biological Chemistry
Sulfiredoxin: a potential therapeutic agent?
Biomedicine and Pharmacotherapy
Deglutathionylation of 2-Cys peroxiredoxin is specifically catalyzed by sulfiredoxin
Journal of Biological Chemistry
Glutaredoxin-1 regulates TRAF6 activation and the IL-1 receptor/TLR4 signalling
Biochemical and Biophysical Research Communications
Characterization of three isoforms of mammalian peroxiredoxin that reduce peroxides in the presence of thioredoxin
Diabetes Research and Clinical Practice
Glutathionylation of adenine nucleotide translocase induced by carbon monoxide prevents mitochondrial membrane permeabilization and apoptosis
Journal of Biological Chemistry
Use and abuse of exogenous H2O2 in studies of signal transduction
Free Radical Biology and Medicine
Nitric oxide signaling: classical, less classical, and nonclassical mechanisms
Free Radical Biology and Medicine
Activation of purified guanylate cyclase by nitric oxide requires heme. Comparison of heme-deficient: heme-reconstituted and heme-containing forms of soluble enzyme from bovine lung
Biochimica et Biophysica Acta
Nox/Duox family of nicotinamide adenine dinucleotide (phosphate) oxidases
Current Opinion in Hematology
Mitochondrial formation of reactive oxygen species
Journal of Physiology
Signaling functions of reactive oxygen species
Biochemistry
Redox signaling: thiol chemistry defines which reactive oxygen and nitrogen species can act as second messengers
American Journal of Physiology: Cell Physiology
The leukotrienes
Medical Biology
Redox sensing: orthogonal control in cell cycle and apoptosis signalling
Journal of Internal Medicine
Aspects of the biological redox chemistry of cysteine: from simple redox responses to sophisticated signalling pathways
Biological Chemistry
Redefining oxidative stress
Antioxidants and Redox Signalling
The role of conjugation reactions in detoxication
Archives of Toxicology
Glutathione transferases
Annual Review of Pharmacology and Toxicology
The role of glutathione in detoxication
Environmental Health Perspectives
Cited by (164)
Clinical physiology and pharmacology of GSTZ1/MAAI
2023, Biochemical PharmacologyRevisiting the scavenging activity of glutathione: Free radicals diversity and reaction mechanisms
2023, Computational and Theoretical ChemistryCellular Red-Ox system in health and disease: The latest update
2023, Biomedicine and PharmacotherapyGlutathione metabolism and regulation in cyanobacteria
2023, Cyanobacteria: Metabolisms to MoleculesPosttranslational modifications and metal stress tolerance in plants
2023, Biostimulants in Alleviation of Metal Toxicity in Plants: Emerging Trends and Opportunities
- ☆
This work was supported by ES005511 from the National Institutes of Health.