Alternative mRNAs arising from trans-splicing code for mitochondrial and cystosolic variants of Echinococcus granulosus thioredoxin glutathione reductase

ECHINOCOCCUS GRANULOSUS

TIOREDOXINA

BIBLIOGRAFIA NACIONAL QUIMICA

Minireviews Reports Classics Reflections Papers of the Week QUICK SEARCHAuthor:Keyword:Year:Vol:Page:GO [Advanced Search][Browse the Archive] Advertisement Advertisement JBC's Young Investigator Award: Learn More Alternative mRNAs Arising from Trans-splicing Code for Mitochondrial and Cytosolic Variants of Echinococcus granulosus Thioredoxin Glutathione Reductase* Astrid Agorio‡§, Cora Chalar¶, Soledad Cardozo‡ and Gustavo Salinas‡‖ + Author Affiliations From the ‡Cátedra de Inmunologı́a, Facultad de Quı́mica/Ciencias, Universidad de la República, Avenida Alfredo Navarro 3051, Piso 2, CP 11.600, Montevideo, Uruguay and ¶Sección Bioquı́mica, Facultad de Ciencias, Universidad de la República, Iguá4225, CP 11.500, Montevideo, Uruguay Next Section Abstract Thioredoxin and glutathione systems are the major thiol-dependent redox systems in animal cells. They transfer via the reversible oxidoreduction of thiols the reducing equivalents of NADPH to numerous substrates and substrate reductases and constitute major defenses against oxidative stress. In this study, we cloned from the helminth parasite Echinococcus granulosus two trans-spliced mRNA variants that encode thioredoxin glutathione reductases (TGR). These variants code for mitochondrial and cytosolic selenocysteine-containing isoforms that possess identical glutaredoxin (Grx) and thioredoxin reductase (TR) domains and differ exclusively in their N termini. Western blot analysis of subcellular fractions with specific anti-TGR antibodies showed that TGR is present in both compartments. The biochemical characterization of the native purified TGR suggests that the Grx and TR domains of the enzyme can function either coupled or independently of each other, because the Grx domain can accept electrons from either TR domains or the glutathione system and the TR domains can transfer electrons to either the fused Grx domain or to E. granulosus thioredoxin. The reversible thiol-disulfide reaction is a central theme in biology. On the one hand, this redox exchange is an efficient mechanism of electron transport. On the other hand, this reaction is a molecular device used by nature as a switch to control protein function and localization through the redox state of critical thiol groups (1). The central role in thiol-disulfide exchange in animal cells is played by the thioredoxin (Trx)1 and glutathione (GSH) systems. Both systems have overlapping and distinct properties and targets but function in a similar way. They transfer via the reversible oxidoreduction of thiols the reducing equivalents of NADPH to numerous substrates and substrate reductases (2). They maintain the cellular redox homeostasis and constitute major defenses against oxidative stress, acting directly by reducing oxidized compounds and proteins or providing reducing equivalents to the hydrogen peroxide reductases present in animal cells, namely thioredoxin peroxidase and glutathione peroxidase (3). Both systems provide electrons to ribonucleotide reductase and control vital cellular processes such as transcription and signal transduction through the redox regulation of kinases, phosphatases, and transcription factors (4-6). The animal thioredoxin system comprises the thioredoxin reductase (TR) and Trx, whereas the glutathione system consists of glutathione reductase (GR), glutathione (GSH), and glutaredoxin (Grx). Trx and Grx are thiol-disulfide oxidoreductases that transfer electrons to various substrates and substrates reductases. GSH, another thiol-based reductant, recycles Grx to its reduced state and reduces other protein and non-protein substrates. TR and GR are pyridine nucleotide-disulfide oxidoreductases that transfer the reducing equivalents of NADPH to oxidized Trx and other substrates and to GSH, respectively. Mammalian and Caenorhabditis elegans TR possess C-terminal electron transfer centers containing redox active cysteine and selenocysteine (Sec) residues (7-10). The selenol group of Sec is a strong nucleophile (11) that confers a profound reductive capacity to the enzyme. The C-terminal redox center is not present in GR. The equivalent redox link in the GSH system is thought to be provided by GSH (10). In mammals, both systems are present in mitochondria and the cytosol. This is because of the existence of genes encoding mitochondrial and cytosolic variants for the proteins from both systems and also because of alternative splicing of single genes (12-17). Recently, mitochondrial and cytosolic variants of DrosophilaTR have also been described to be derived from alternative splicing of a single gene (18). Recently, a more complex machine exhibiting specificity for both thioredoxin and glutathione systems has been characterized in mammals (19). This enzyme termed thioredoxin glutathione reductase (TGR) is a selenoprotein oxidoreductase that possesses thioredoxin reductase, glutathione reductase, and glutaredoxin activities, achieving this broad substrate specificity by a fusion of TR and Grx domains. This enzyme has been reported to be present in the microsomal fraction of testis cells from mice. A multifunctional TGR has also been reported inSchistosoma mansoni, a platyhelminth organism (20), indicating that this fusion domain has been evolutionarily conserved. In this article, we report the cloning and characterization of a thioredoxin glutathione reductase of the larval stage ofEchinococcus granulosus, a cestode parasite that dwells in the liver and lungs of human and cattle hosts. The results indicate that the enzyme possesses glutathione reductase, thioredoxin reductase, and glutaredoxin activities and suggest that there is domain communication between the C-terminal Cys-Sec redox center and the N-terminal glutaredoxin domain. In addition, we provide conclusive evidence that mitochondrial and cytosolic variants of TGR are generated from a single TGR gene

The Journal of Biological Chemistry v. 278, no. 15, 2003. -- p. 12920-12928

The American Society for Biochemistry and Molecular Biology, Inc.

2003

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DOI: 10.1074/jbc.M209266200