GSH (L-γ-glutamyl-L-cysteinyl-glycine) is a linear tripeptide (M.W. 307.4 g mol⁻¹) formed from the amino acids glycine, cysteine, and glutamate. In solution, GSH possess a net negative charge of −1 at physiological pH, where the l-glutamic acid predominantly exists in its zwitterionic form, while the carboxyl group of the glycine fragment prefers to be deprotonated. GSH, glutathione.

Fates of GSH during cell death progression. GSH depletion during cell death can occur by distinct mechanisms. (1) Upon oxidative stress, GSH is used for the scavenging of peroxides by GPxs, which generate GSSG as a byproduct. GSSG can be reduced back to GSH by the GR/NADPH system. (2) GSH loss also occurs via its extrusion across the plasma membrane by the activation of GSH transporters or pumps (GSH-T). (3) GSH-Ts also mediate GSSG efflux and transport of GSH-conjugates (GS-XN) generated by xenobiotics in order to avoid deleterious effects of the accumulation of these toxins. (4) GSH depletion might also be associated with the impairment of GSH de novo synthesis as demonstrated by the impairment of the cysteine uptake transporters (EAAC1 in neurons) and the degradation of GCL by caspases. (5) Alterations in GSH/GSSG balance during apoptosis have been correlated with alterations in PSSG levels. In addition, other oxidative forms of GSH such as GSNO might also be formed by the direct interaction of GSH with distinct ROS/RNS. GCL, glutamate-cysteine ligase; GR, glutathione reductase; GSH, glutathione; GSSG, glutathione disulfide; GSNO, S-nitrosoglutathione; NADPH, nicotinamide adenine dinucleotide phosphate; PSSG, protein glutathionylated; ROS/RNS, reactive nitrogen species/reactive oxygen species.

Compartmentalization of the GSH/GSSG redox couple. GSH is produced intracellularly and is found 70%–90% freely distributed in the cytosol, but also compartmentalized in mitochondria, nuclear matrix, and ER. Specific transport mechanisms maintain compartmentalized GSH/GSSG homeostasis. GSH diffuses through MOM via porin channels (not depicted here), and translocates through the IMM via DIC or OGC exchangers. In the nucleus, GSH is considered to diffuse freely through the nuclear pore. Protein-dependent facilitated diffusion is thought to mediate GSH permeation in the ER, but the molecular identify of the mechanism(s) involved remains unknown. Within the ER, GSH exits largely as GSSG due to its oxidation. It has been proposed that GSSG could be secreted via the secretory pathway for its recycle. A variety of protein transporters have been reported to act as plasma membrane GSH transporters (GSH-T), but their role in GSH depletion during cell death progression is still unclear. Values indicate redox potential for GSH/GSSG (mV), % of compartmentalized GSH with respect to total cellular levels, concentration of GSH (mM), and GSH/GSSG ratio for each subcellular compartment. Values were taken from (, , , ). IMM, inner mitochondrial membrane; DIC, dicarboxylate carrier; OGC, 2-oxoglutarate transporters.

Plasma membrane GSH efflux pumps. Distinct candidates have been proposed to act as GSH transporters. The MRPs act as ATP-dependent cotransporters of GSH (coupled to the extrusion of an OA⁻), GSSG, and GSH conjugates. MRP1 can transport GSH alone, but this requires its stimulation by xenobiotics. The OATPs were initially proposed to act as the GSH/OA⁻ exchanger, where GSH efflux is thought to be driven by its electrochemical gradient across the plasma membrane, and stimulated by the presence of extracellular OA⁻. Other proposed candidates for GSH efflux are the members of the ABC family of transporter CFTR and BCRP/ABCG2, hemichannel connexins (CX), and RLIP76. Energy dependency of GSH transport by BCRP/ABCG2 has not yet been confirmed. ABC, ATP-binding cassette; MRP, multidrug resistance protein; OA⁻, organic anion; OATP, organic anion transporting polypeptide; RLIP76 (RALBP1), Ral-binding, Rho/Rac-GAP and Ral effector.

Molecular mechanisms involved in the regulation of apoptosis by GSH. GSH depletion regulates cell death progression by apoptosis through a variety of mechanisms. GSH depletion triggers the permeability transition pore of the mitochondria, the pro-apoptotic function of released Cyt C, the formation of the apoptosome, and the activation of executioner caspases. Furthermore, GSH depletion precedes oxidative stress and is necessary for ROS/RNS formation. Alterations in GSH, GSSG, GSNO, and ROS/RNS homeostasis can modify the levels of PSSG/PSNO residues. Aggregation of death receptors and caspase activation has been demonstrated to be regulated by protein glutathionylation and nitros(yl)ation. Cyt C, cytochrome C; BH3, Bcl-2 homology 3 ; Bcl-2, B-cell lymphoma 2; Bid, BH3 interacting-domain death agonist; FADD, Fas-associated death domain; GSNO, S-nitroglutathione; PSSG, protein glutathionylation; PSNO, protein nitros(yl)ation; ROS/RNS, reactive oxygen and nitrogen species.

Redox alterations induced by GSH/GSSG transport. (1) Several protein transporters have been proposed to mediate GSH/GSSG transport, which in fact, can significantly impact cellular redox balance. (2) GSH levels maintain a reduced intracellular environment, even under normal conditions as evidenced by observations that by itself GSH depletion induces oxidative stress. GSH directly scavenges ROS/RNS or enzymatically, through the GPx/GR/NADPH/G6PD system. Thus, GSH efflux sensitizes cells to oxidative stress, while GSSG transport can serve as a protective mechanism. (3) Changes in the GSH:GSSG ratio directly result in alterations of oxidative post-translational modifications in protein thiols (PSH). (4) Both GSH and GSSG have the ability to promote PSSG formation via (a) GSSG reaction with PSH, (b) GSH reaction with PSOHs, the most commonly accepted mechanism, and (c) Grx-mediated transfer of thiyl radicals (GS•) to PSH residues. (5) PSSGs are known to regulate enzyme function and activity (redox signaling) and protect cysteines from irreversible oxidation to PSO₂H and PSO₃H residues, and subsequent degradation. Gpx, glutathione peroxidase; GR, glutathione reductase; G6PD, glucose-6-phosphate dehydrogenase; Grx, glutaredoxin; NADPH, nicotinamide adenine dinucleotide phosphate; PSOH, protein sulfenic acid; PSO₃H, protein sulfonic acids; PSO₂H, protein sulfinic acids.