Thiol-based redox regulation has emerged as the prevalent mechanism for redox regulation of cellular processes. Its major component, the thioredoxin (Trx) system, is present in all organisms. Thioredoxin reductases (TRs) control the redox state of Trxs, which in turn regulate cellular processes by controlling the redox state of Trxs. TRs and Trxs have been implicated in cancer development, apoptosis, signal transduction and many other processes, but the pathways and mechanisms of thiol-based regulation are poorly understood. Mammals have three TRs: cytosolic TR1 (Txnrd1), mitochondrial TR3 (Txnrd2) and thioredoxin/glutathione reductase TGR (Txnrd3). All three contain a catalytic selenocysteine in the C-terminal region and, therefore, the mammalian Trx system is dependent on dietary selenium. In addition, each TR has several isoforms that differ by their N-terminal sequences. Among TR substrates, previous studies focused on Trx1 and few other thiol oxidoreductases (i.e., proteins that use catalytic redox Cys). However, there are numerous other Trx-like proteins and thiol oxidoreductases, and the overall composition of this system and identity of proteins regulated by thiol oxidoreductases is not known. This is important because thiol oxidoreductases provide a framework for redox regulation of cellular processes by targeting redox Cys residues in proteins. Previously, we defined the basic properties of TRs and their roles in various biological processes, organs and organisms. Additional data revealed successful application of mechanism-based approaches to identify substrates of TRs and thiol oxidoreductases. Computational approaches were also developed that allow identification of entire sets of thiol oxidoreductases in organisms. We now propose to use this foundation to characterize isoforms of TRs, their thiol oxidoreductase substrates and protein targets of thiol oxidoreductases to define the major pathway of thiol-based redox regulation in mammals. To address this challenge, we propose the following specific aims: (1) Characterize regulation of TR function and redox homeostasis by heterodimerization and position-specific Sec insertion. (2) Determine specific substrates of mammalian TR isoforms. (3) Define a mammalian set of thiol oxidoreductases and their protein targets. A combination of computational, proteomic, cell biology, and animal model approaches will be used to address these questions, and the data will be used to define the network of thiol-based redox regulation of protein function in mammals.

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A redox system composed of thioredoxin reductase and thioredoxin is the main regulator of the redox state of cysteines in proteins, whereby controlling cellular redox homeostasis and major metabolic processes. This system has been implicated in cell signaling, apoptosis, cancer development, and many other physiological and pathophysiological processes. Using computational, proteomic, cell biology, and animal model approaches, we propose to determine functions of thioredoxin reductase isoforms, their thiol oxidoreductase substrates and identities of downstream targets to define a skeleton of thiol-based redox regulation in mammals.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Method to Extend Research in Time (MERIT) Award (R37)
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Special Emphasis Panel (ZRG1-EMNR-H (02))
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Anderson, Vernon
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Brigham and Women's Hospital
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Malinouski, Mikalai; Hasan, Nesrin M; Zhang, Yan et al. (2014) Genome-wide RNAi ionomics screen reveals new genes and regulation of human trace element metabolism. Nat Commun 5:3301
Otero, LucĂ­a; Romanelli-Cedrez, Laura; Turanov, Anton A et al. (2014) Adjustments, extinction, and remains of selenocysteine incorporation machinery in the nematode lineage. RNA 20:1023-34
Labunskyy, Vyacheslav M; Suzuki, Yo; Hanly, Timothy J et al. (2014) The insertion Green Monster (iGM) method for expression of multiple exogenous genes in yeast. G3 (Bethesda) 4:1183-91
Labunskyy, Vyacheslav M; Gerashchenko, Maxim V; Delaney, Joe R et al. (2014) Lifespan extension conferred by endoplasmic reticulum secretory pathway deficiency requires induction of the unfolded protein response. PLoS Genet 10:e1004019
Kaya, Alaattin; Lobanov, Alexei V; Gerashchenko, Maxim V et al. (2014) Thiol peroxidase deficiency leads to increased mutational load and decreased fitness in Saccharomyces cerevisiae. Genetics 198:905-17
Gerashchenko, Maxim V; Gladyshev, Vadim N (2014) Translation inhibitors cause abnormalities in ribosome profiling experiments. Nucleic Acids Res 42:e134
Labunskyy, Vyacheslav M; Hatfield, Dolph L; Gladyshev, Vadim N (2014) Selenoproteins: molecular pathways and physiological roles. Physiol Rev 94:739-77
Turanov, Anton A; Lobanov, Alexei V; Hatfield, Dolph L et al. (2013) UGA codon position-dependent incorporation of selenocysteine into mammalian selenoproteins. Nucleic Acids Res 41:6952-9
Hondal, Robert J; Marino, Stefano M; Gladyshev, Vadim N (2013) Selenocysteine in thiol/disulfide-like exchange reactions. Antioxid Redox Signal 18:1675-89
Tobe, Ryuta; Naranjo-Suarez, Salvador; Everley, Robert A et al. (2013) High error rates in selenocysteine insertion in mammalian cells treated with the antibiotic doxycycline, chloramphenicol, or geneticin. J Biol Chem 288:14709-15

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