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Review
, 18 (10), 1165-207

Thioredoxin and Thioredoxin Target Proteins: From Molecular Mechanisms to Functional Significance

Affiliations
Review

Thioredoxin and Thioredoxin Target Proteins: From Molecular Mechanisms to Functional Significance

Samuel Lee et al. Antioxid Redox Signal.

Abstract

The thioredoxin (Trx) system is one of the central antioxidant systems in mammalian cells, maintaining a reducing environment by catalyzing electron flux from nicotinamide adenine dinucleotide phosphate through Trx reductase to Trx, which reduces its target proteins using highly conserved thiol groups. While the importance of protecting cells from the detrimental effects of reactive oxygen species is clear, decades of research in this field revealed that there is a network of redox-sensitive proteins forming redox-dependent signaling pathways that are crucial for fundamental cellular processes, including metabolism, proliferation, differentiation, migration, and apoptosis. Trx participates in signaling pathways interacting with different proteins to control their dynamic regulation of structure and function. In this review, we focus on Trx target proteins that are involved in redox-dependent signaling pathways. Specifically, Trx-dependent reductive enzymes that participate in classical redox reactions and redox-sensitive signaling molecules are discussed in greater detail. The latter are extensively discussed, as ongoing research unveils more and more details about the complex signaling networks of Trx-sensitive signaling molecules such as apoptosis signal-regulating kinase 1, Trx interacting protein, and phosphatase and tensin homolog, thus highlighting the potential direct and indirect impact of their redox-dependent interaction with Trx. Overall, the findings that are described here illustrate the importance and complexity of Trx-dependent, redox-sensitive signaling in the cell. Our increasing understanding of the components and mechanisms of these signaling pathways could lead to the identification of new potential targets for the treatment of diseases, including cancer and diabetes.

Figures

FIG. 1.
FIG. 1.
Three-dimensional structure of human thioredoxin (Trx). The basic thioredoxin fold consists of four β-beta strands surrounded by three α-helices. Trx has an additional α-helix and β-beta strand at the N-terminus. The catalytically active cysteine residues at position 32 and 35 are highlighted. The image was generated using the PyMOL Molecular Graphics System (Schrödinger, LLC, Portland, NY) based on PDB ID: 1TRW.
FIG. 2.
FIG. 2.
Redox cascade of the Trx system. Reduced nicotinamide adenine dinucleotide phosphate (NADPH)+H+ is generated by the pentose phosphate pathway. NADPH+H+ reduces oxidized Trx reductase (TrxR), which regenerates the pool of reduced Trx. Reduced Trx contributes to maintaining a reducing environment for a number of different proteins.
FIG. 3.
FIG. 3.
Catalytic reaction of Trx reductase. NADPH+H+ reduces enzyme-bound flavine adenine dinucleotide (FAD) in one subunit. The reducing equivalents are transferred to the -CVNVGC- active site motif of the same subunit forming a dithiol motif (not shown). This dithiol motif reduces the C-terminal selenenyl sulfide motif (not shown) of the other subunit of the homodimer forming a selenolthiol motif. The reduced selenolthiol motif can reduce the substrates of TrxR, including the active site disulfide of Trx.
FIG. 4.
FIG. 4.
Trx target proteins. Oxidized Trx is reduced by TrxR to maintain a pool of reduced Trx. Trx reduces peroxiredoxin (Prx), ribonucleotide reductase (RNR), and methionine sulfoxide reductase (Msr). These reductive enzymes catalyze the reduction of peroxides, ribonucleotides, and methionine sulfoxides, respectively. Trx also directly interacts with redox-sensitive molecules, such as apoptosis signal-regulated kinase 1 (ASK1), thioredoxin interacting protein (Txnip), and phosphatase and tensin homolog (PTEN). Redox-sensitive molecules modulate different cellular processes, including development, proliferation, migration, apoptosis, inflammation, and metabolism.
FIG. 5.
FIG. 5.
Catalytic reaction of 2-Cys Prxs. Hydrogen peroxide (H2O2) oxidizes the thiol residue of the N-terminal cysteine of Prx I–IV. The resulting sulfenic acid reacts with the thiol residue of the C-terminal cysteine of a second Prx to form an intermolecular disulfide bond. In this reaction, H2O2 is reduced to H2O, while Prx is oxidized into a homodimer. Trx catalyzes the reduction of oxidized Prx.
FIG. 6.
FIG. 6.
Mitogen-activated protein kinase (MAPK) signaling cascade. The MAPK pathway consists of sequential MAP kinase kinase kinase (MAPKKK), MAP kinase kinase (MAPKK), and MAPK activations. Extracellular stimuli and stress, such as growth factors, reactive oxygen species (ROS), and ultraviolet (UV) irradiation, are detected by MAPKKK. The subsequent activation of MAPKK and MAPK regulates downstream targets to control diverse cellular activities, including stress response, apoptosis, cell-cycle arrest, cell survival, innate immunity, proliferation, and differentiation.
FIG. 7.
FIG. 7.
Domain and three-dimensional structure of human ASK1. (A) Human apoptosis signal-regulating kinase 1 (ASK1) contains a serine (Ser)/threonine (Thr) kinase domain. Phosphorylation of a specific Thr residue in the kinase domain, T838, is required for ASK1 activation. ASK1 also possesses a phosphoserine motif that is recognized by the 14-3-3 proteins which regulate the activity of ASK1. The N-terminal coiled coil (NCC) and C-terminal coiled coil (CCC) domains are important for binding other regulatory proteins, including Trx and tumor necrosis factor receptor-associated factor (TRAF) 2 and TRAF6. (B) In the absence of stress, ASK1 forms a homo-oligomer that is stabilized by the CCC domain. ASK1 signalosome that is required for the proper ASK1 regulation includes this homo-oligomer, as well as other ASK1 regulating proteins such as Trx and TRAF 2/6. The activation of ASK1 occurs through the phosphorylation of T838. The image was generated using PyMOL Molecular Graphics System (Schrödinger, LLC) based on PDB ID: C2LQ.
FIG. 8.
FIG. 8.
Activation of ASK1 through signalosome formation. Regulatory proteins that bind to ASK1 control its activity. Once reduced, Trx binds to ASK1, ASK1 activation is suppressed. However, in the presence of ROS or other types of stress, Trx is oxidized and dissociates from the signalosome. In the presence of Txnip, a disulfide is formed between Txnip and Txn, inducing the dissociation of Txn from ASK1. Phosphorylation of ASK1 and the recruitment of TRAF2/6 proteins activate ASK1.
FIG. 9.
FIG. 9.
Phylogenic tree of α- and β-arrestins. The evolutionary history was inferred using the Neighbor-Joining method. The optimal tree with the sum of branch length=5.29379310 is shown. The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the Poisson correction method and are in the units of the number of amino-acid substitutions per site. The analysis involved 10 amino-acid sequences. All positions containing gaps and missing data were eliminated. There were a total of 288 positions in the final dataset. Evolutionary analyses were conducted in MEGA5 (299). α-arrestins: ARRDC1–5=arrestin domain containing 1–5; β-arrestins: SAG=visual arrestin=arrestin 1; ARRB1=β-arrestin 1=arrestin 2; ARRB2=β-arrestin 2=arrestin 3; ARR3=X-arrestin=arrestin 4.
FIG. 10.
FIG. 10.
Domain structure of human Txnip. Txnip shares the common domain structure of arrestin proteins with an N-terminal and a C-terminal arrestin domain. The latter contains a cysteine at position 247, which is required for the interaction with Trx. At the C-terminal, Txnip has two PPXY motifs, which are known to interact with WW domains.
FIG. 11.
FIG. 11.
Proposed Txnip—Trx interaction. (A) Txnip reacts with reduced Trx and forms an intermolecular disulfide at cysteine 247. (B) No disulfide exchange reaction is possible between Txnip and oxidized Trx.
FIG. 12.
FIG. 12.
Transcriptional regulation of Txnip. (A) Transcription of TXNIP is induced by glucose through a number of different transcription factors, including Krüppel-like factor 6 (KLF6), which binds to a GC box (GC), carbohydrate response element-binding protein (ChREBP), and MondoA:Max-like protein X (Mlx), which bind to carbohydrate response elements (ChoREs), forkhead box O 1 (FOXO1), which binds to a FOXO bindings site (FOXO), and heterotrimeric nuclear factor Y (NFY), which binds to a CCAAT box (CCAAT) and an inverted CCAAT box (ATTGG). Other inducers of Txnip expression are lactic acidosis, hypoxia, inhibition of oxidative phosphorylation (OXPHOS), and adenosine-containing molecules (NADH/ATP) through MondoA:Mlx, glucocorticoids (G) through translocation of the glucocorticoid receptor (GR) and binding to a glucocorticoid response element (GRE), heat shock through heat shock factor 1 (HSF1) and a heat shock element (HSE), peroxisome proliferator-activated receptor gamma (PPARγ) activation through PPARγ:retinoid X receptor (RXR) binding to a PPARγ response element (PPRE), synthetic retinoid CD437 through E-twenty six 1 (ETS) and an ETS1-binding sequence (EBS), and suberoylanilide hydroxamic acid (SAHA) through the disinhibition of transcription by the inhibition of histone deacetylase 1 (HDAC1), which is associated with the Txnip promoter through a complex of RET finger protein (RFP) and NFY. PPARα activation leads to inhibition of the heterodimeric activator protein 1 (AP1) complex that consists of c-Fos and c-Jun, resulting in suppression of Txnip expression. (B) Epigenetic suppression of Txnip expression includes histone deacetylation mediated by HDAC1, methylation of CpG sites (CG) by DNA methyltransferase (DNMT), and trimethylation of histone H3 at K27 by polycomb repressive complex 2 (PRC2). Treatment with decitabine, SAHA or 3-deazaneplanocin A (DZNep) results in an increased expression of Txnip.
FIG. 13.
FIG. 13.
Mechanism of inhibition of proliferation by Txnip. Txnip forms a transcriptional repressor complex with Fanconi anemia zinc finger (FAZF), promyelocytic leukemia zinc finger (PLZF), and HDAC1. This complex binds to a PLZF-response element in the cyclin A2 promoter and suppresses cyclin A2 expression. Cyclin A2 activates cyclin-dependent kinase 2 (CDK2), which is required for G1/S transition in the cell cycle. Txnip interacts with HDAC1 and HDAC3, which bind to NF-κB p50 and p65 and leads to transcriptional repression at the NF-κB binding site. Jun activation-domain binding protein 1 (JAB1) induces the translocation of CDK2/4 inhibitor p27kip1 from the nucleus to the cytosol where it is degraded. Txnip interacts with JAB1 and blocks JAB1-mediated translocation of p27kip1 to the cytosol. Txnip increases protein expression of p16Ink4a, an inhibitor of CDK4/6, resulting in decreased G1/S cell-cycle transition and decreased proliferation. Regulated in development and DNA damage responses 1 (Redd1) protein is a negative regulator of mammalian target of rapamycin (mTOR) acitivity and proliferation. Txnip interacts with Redd1 and protects it from proteasomal degradation.
FIG. 14.
FIG. 14.
Proposed mechanism of regulation of adipogenesis by Txnip. (A) Wild-type Txnip binds to Trx and is protected against ubiquitination. Txnip inhibits PPARγ activity, which results in decreased adipogenesis. There might be other proteins that are potentially involved in the inhibition of PPARγ activity. (B) Mutation of Txnip cysteine 247 to Ser abrogates binding to Trx and allows E3 ubiquitin ligase Itch to bind Txnip at its PPXY domains, resulting in the ubiquitination and degradation of Txnip. Disinhibition of PPARγ leads to increased adipogenesis. The recruitment of Itch by Txnip might lead to the ubiquitination of other proteins involved in the regulation of PPARγ activity.
FIG. 15.
FIG. 15.
Domain structure of human PTEN. PTEN contains a lipid/protein phosphatase domain in the N-terminal region. In the C-terminal domain, PTEN is composed of a C2 domain, two proline, glutamate, serine, threonine (PEST) domains, and a PDZ domain. C2 domain allows PTEN to interact with various lipids. PEST domains are rich in proline, glutamate, Ser, and Thr residues. PEST and PDZ domains may be important for both PTEN stabilization and their interactions with other proteins.
FIG. 16.
FIG. 16.
Regulation of PTEN by Trx and Txnip. Active PTEN is represented by reduced C212 of the C2 domain of PTEN. On formation of a disulfide between C212 of PTEN and C32 of thioredoxin, PTEN becomes inactive, resulting in cell survival and proliferation. However, the inactivation of PTEN by thioredoxin can be reversed in the presence of Txnip. A disulfide that is formed between the C247 of Txnip and the C32 of thioredoxin allows for the reactivation of PTEN.

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