Department of Selenoprotein Research


Prof. Elias Arnér MD PhD

Head of Department
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Dr. Marcell Cserhalmi

Laboratory manager
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Anikó Keszőcze

Assistant administrative
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Dr. Attila Andor

Biologist
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Dr. Mahendravarman Mohanraj

Biologist
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Dr. Katalin Úri

Biologist
CV



Staff of the Department

Prof. Dr. Elias Arnér MD PhD Head of Department
contact: elias.arner@ki.se 
tel: +46 - 8 - 5248 69 83 (KI office) 
 
 
Dr. Marcell Cserhalmi
Laboratory manager
contact: cserhalmi.marcell@oncol.hu
tel: +36-1-224-8600 / 1353

 

Dr. Katalin Úri
biologist
E-mail: uri.katalin@oncol.hu
tel.: +36-1-224-8600 / 1352

 
Anikó Keszőcze
Laboratory coordinator
e-mail: keszocze.aniko@oncol.hu
tel: +36-1-224-8600 / 3774


Dr. Attila Andor
Biologist
e-mail: andor.attila@oncol.hu tel: +36-1-224-8600 / 1353
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Dr. Mahendravarman Mohanraj
Biologist
tel: +36-1-224-8600 / 3492

Research of the Department (Department of Selenoprotein Research at NIO)

This department at NIO was newly founded in 2020 as a pure research department, dedicated to research on the function and importance of selenoproteins in health and disease. For its establishment, NIO recruited Prof. Elias Arnér from Karolinska Institutet in Stockholm, Sweden, who is a world authority in the field of selenoprotein research. Prof. Arnér concomitantly maintains his position at Karolinska Institutet while he also has a part-time affiliation at NIO as Head of the Department of Selenoprotein Research. The activities at the Department are thereby closely linked to the research conducted by Prof. Arnér in Stockholm, with exchange of staff and research ideas between the two sites.  For further information on the activities of the Arnér group in Stockholm, see https://ki.se/en/mbb/elias-arner-research-group For publications by Prof. Arnér, see https://www.ncbi.nlm.nih.gov/myncbi/elias.arner.1/bibliography/public/.

Schematic drawing of the selenium-containing amino acid Selenocystine (Sec) in comparison to the more common sulfur-containing amino acid Cysteine (Cys). Most functions of selenium – an essential trace element for humans – are carried out by selenoproteins, containing Sec as their defining entity. 

For more information see 

Arnér ES. Selenoproteins-What unique properties can arise with selenocysteine in place of cysteine? Exp Cell Res. 2010;316:1296–1303. doi:10.1016/j.yexcr.2010.02.032

 

Overriding aim and significance of our research 

Our major aim is to increase the understanding of selenoprotein functions and their significance for health and disease, especially cancer, with the purpose of improving medical therapeutics that target redox perturbations in disease and developing new applications based upon recombinant selenoproteins. Specifically, we aim to answer the following questions:     How can selenoproteins be produced with high efficiency, to facilitate in-depth studies of their molecular mechanisms and functions?     What are the roles of specific selenoproteins in cancer and how can their unique biochemical features be utilized to improve cancer treatment?

 

SIGNIFICANCE 

Our studies are expected to yield important insights into functions of Sec in selenoproteins, having major importance in several aspects of health and disease. Specifically, our results provide mechanistic insights into the roles of TrxR1 and GPx4 as targets for anticancer therapy, enabling the development of more efficient and specific anticancer therapy protocols. Our novel biotechnological applications based upon recombinant selenoproteins are of potential importance in several medical fields, specifically personalized cancer diagnostics using functional imaging and novel modalities for cancer treatment. Collectively, these studies lead to increased knowledge of selenoprotein function, with insights into molecular mechanisms geared towards improved cancer care.

 

Selenoproteins

Our studies focus on selenoproteins, i.e. proteins carrying the rare selenium-containing selenocysteine (Sec) residue (1-3). Because Sec is significantly more reactive than its common sulfur-containing Cys homolog, selenoproteins have unique features with high chemical reactivity and facilitated catalysis of redox reactions (1). There are exactly 25 genes in the human genome encoding selenoproteins, including enzymes such as thyroid hormone deiodinases, thioredoxin reductases (TrxRs) and glutathione peroxidases (GPXs) (4). Selenium deficiency results in insufficient selenoprotein synthesis with cardiomyopathy, cancer and male infertility (2). Knockout mice deficient in tRNA for Sec die at implantation, similar to the embryonic death of mice lacking thioredoxin (Trx) or TrxRs. Thus, TrxR isoforms are essential for life, or at least for embryonic development. TrxRs have a catalytic Sec residue at their C-terminal ends and catalyze several forms of redox reactions, the understanding of which has been a main focus of our studies for the last two decades (5-8). It should be underscored that the whole human Trx system, with a wide range of functions, is fully selenium dependent because of TrxRs being selenoproteins. 

 

Synthesis of selenoproteins involves a natural expansion of the genetic code

Insertion of Sec occurs co-translationally at dedicated UGA codons. Because UGA is normally a stop codon, expression of selenoproteins necessitates a natural expansion of the genetic code. This occurs through Sec-specialized translation factors interacting with an mRNA structure called a SECIS (Sec Insertion Sequence). SECIS features differ between bacteria and mammals and make direct expression of mammalian selenoproteins in E. coli impossible. Mainly due to a lack of selenoprotein supply, few selenoproteins have yet been obtained in larger amounts for in-depth studies. However, in 1999 we expressed mammalian TrxR1 in E. coli by engineering a bacterial-type SECIS element, which was the first time a mammalian selenoprotein was produced in bacteria (9). Inspired by the success of that approach, we developed a Sel-tag for recombinant proteins mimicking the C-terminus of TrxR1 (-Gly-Cys-Sec-Gly). The high chemical reactivity of Sec in this Sel-tag could be used as a chemical "handle" for site-directed targeting, including radiolabeling with positron emitters for PET imaging. Our first study of the Sel-tag was published in the launch issue of Nature Methods (10) and further described in the first volume of Nature Protocols (11). We have since developed the Sel-tag for diagnostics (12-14) and continue to refine this methodology, which is one line of our research. The methods that we have developed for selenoprotein production puts us in a unique position worldwide for studies of selenoprotein functions.

Scheme of the features of selenoprotein-encoding mRNAs and the differences between those of bacteria compared to mammals. 

For more information see 

Arnér ES. Recombinant expression of mammalian selenocysteine-containing thioredoxin reductase and other selenoproteins in Escherichia coli. Methods Enzymol. 2002;347:226–235. doi:10.1016/s0076-6879(02)47022-x 

Cheng Q, Arnér ES. Selenocysteine Insertion at a Predefined UAG Codon in a Release Factor 1 (RF1)-depleted Escherichia coli Host Strain Bypasses Species Barriers in Recombinant Selenoprotein Translation. J Biol Chem. 2017;292(13):5476–5487. doi:10.1074/jbc.M117.776310

 

TrxR1 and GPX4 are selenoproteins that are likely to serve as promising anticancer drug targets

 Several electrophilic anticancer drugs target the reactive Sec residue of TrxR1, which explains some of their efficacy (15,16). We also found that selenium compromised forms of TrxR1 induce cell death by a gain of function in the form of SecTRAPs (Selenium compromised thioredoxin reductase-derived apoptotic proteins). Formation of SecTRAPs may accelerate both efficacy and side effects of electrophilic drugs used in cancer therapy (17,18). Recently we performed a large screening effort to find novel compounds targeting TrxR1 and producing SecTRAPs, finding that such compounds indeed also function as potent anticancer agents, at least in mouse models (19). GPx4, essential during development and for certain interneurons in mice (20), is also a potentially important selenoprotein anticancer drug target. To further understand and employ these selenoproteins for improvement of anticancer drug protocols is another important aspect of our research. 


Many clinically used anticancer drugs target the Sec of TrxR1. 

For more information see:

Busker S, Qian W, Haraldsson M, et al. Irreversible TrxR1 inhibitors block STAT3 activity and induce cancer cell death. Sci Adv. 2020;6(12):eaax7945. Published 2020 Mar 20. doi:10.1126/sciadv.aax7945

Arnér ESJ. Targeting the Selenoprotein Thioredoxin Reductase 1 for Anticancer Therapy. Adv Cancer Res. 2017;136:139–151. doi:10.1016/bs.acr.2017.07.005

Eriksson SE, Prast-Nielsen S, Flaberg E, Szekely L, Arnér ES. High levels of thioredoxin reductase 1 modulate drug-specific cytotoxic efficacy. Free Radic Biol Med. 2009;47(11):1661–1671. doi:10.1016/j.freeradbiomed.2009.09.016

 

Recombinant production of selenoproteins

Recently, after almost two decades of efforts, we finally succeeded in developing a method for production in E. coli of recombinant selenoproteins containing internal Sec residues, which is much more challenging than expressing TrxRs or a Sel-tag, with Sec located close to the C-terminus. We found that using a release factor 1-depleted host strain with tailored insertion of Sec at a UAG codon (instead of UGA), we are able to produce selenoproteins with internal Sec residues without the use of a SECIS element, which we demonstrated with production of human GPX1 (21). This was a major breakthrough for us, as the technique opens up the power of recombinant production for virtually any kind of selenoprotein. To build upon the potential of this powerful method is hence a major aspect of our continued research. We will now, for the first time, be able to compare the two major GPX isoenzymes, GPX1 and GPX4 (which we recently succeeded in producing in the form of active selenoprotein enzymes), side-by-side in terms of biochemical characteristics. GPX1 is believed to be important in mainly reducing H2O2 in the cytosol, while GPX4 adheres to cellular membranes and is known to reduce phospholipid hydroperoxides and protect cells from the iron-triggered and lipid peroxide-dependent ferroptotic type of cell death. However, shared or overlapping activities between the two enzymes have been insufficiently studied. After our initial characterizations of the two enzymes, we subsequently aim to screen both isoenzymes for selective inhibitors, following a similar path in collaboration with the screening facility of NIH at Bethesda, USA, as we did in our discovery of novel TrxR1 inhibitors (19). 

 

Anticancer therapies targeting selenoproteins

Recently we published novel inhibitors of TrxR1 having anticancer properties (19). To further study the anticancer potentials of these inhibitors is one important aspect of our studies. With targeting of GPX4 likely being a promising rationale for cancer therapy, as the enzyme can protect cancer cells from ferroptosis, specific inhibitors of GPX4 would likely complement the targeting of TrxR1 in cancer therapy, and overall expand the therapeutic potential of selenoprotein drug targeting. At NIO we have good possibilities to take our inhibitors to clinical applications, considering the excellent oncology expertise and facilities available at NIO.

 

References

1.    Arnér, E. S. J. (2010) Selenoproteins-What unique properties can arise with selenocysteine in place of cysteine? Exp Cell Res 316, 1296-1303 2.    Rayman, M. P. (2000) The importance of selenium to human health. Lancet 356, 233-241 3.    Johansson, L., Gafvelin, G., and Arnér, E. S. J. (2005) Selenocysteine in proteins — properties and biotechnological use. Biochim Biophys Acta 1726, 1-13 4.    Kryukov, G. V., Castellano, S., Novoselov, S. V., Lobanov, A. V., Zehtab, O., Guigo, R., and Gladyshev, V. N. (2003) Characterization of mammalian selenoproteomes. Science 300, 1439-1443 5.    Cheng, Q., Sandalova, T., Lindqvist, Y., and Arnér, E. S. J. (2009) Crystal structure and catalysis of the selenoprotein thioredoxin reductase 1. J Biol Chem 284, 3998-4008 6.    Gromer, S., Johansson, L., Bauer, H., Arscott, L. D., Rauch, S., Ballou, D. P., Williams Jr, C. H., Schirmer, R. H., and Arnér, E. S. J. (2003) Active sites of thioredoxin reductases — Why selenoproteins? Proc. Nat'l Acad. Sci. U.S.A. 100, 12618-12623 7.    Zhong, L., Arnér, E. S. J., Ljung, J., Åslund, F., and Holmgren, A. (1998) Rat and calf thioredoxin reductase are homologous to glutathione reductase with a carboxyl-terminal elongation containing a conserved catalytically active penultimate selenocysteine residue. J. Biol. Chem. 273, 8581-8591 8.    Nordberg, J., Zhong, L., Holmgren, A., and Arnér, E. S. J. (1998) Mammalian thioredoxin reductase  is irreversibly inhibited by dinitrohalobenzenes by alkylation of both the redox active selenocysteine and its neighboring cysteine residue. J. Biol. Chem. 273, 10835-10842 9.    Arnér, E. S. J., Sarioglu, H., Lottspeich, F., Holmgren, A., and Böck, A. (1999) High-level expression in Escherichia coli of selenocysteine-containing rat thioredoxin reductase utilizing gene fusions with engineered bacterial-type SECIS elements and co-expression with the selA, selB and selC genes. J. Mol. Biol. 292, 1003-1016 10.    Johansson, L., Chen, C., Thorell, J.-O., Fredriksson, A., Stone-Elander, S., Gafvelin, G., and Arnér, E. S. J. (2004) Exploiting the 21st amino acid - purifying and labeling proteins by selenolate targeting. Nat Methods 1, 61-66 11.    Cheng, Q., Stone-Elander, S., and Arnér, E. S. J. (2006) Tagging recombinant proteins with a Sel-tag for purification, labeling with electrophilic compounds or radiolabeling with carbon-11. Nat Protocols 1, 604-613 12.    Cheng, Q., Lu, L., Grafstrom, J., Olofsson, M. H., Thorell, J. O., Samen, E., Johansson, K., Ahlzen, H. S., Linder, S., Arner, E. S., and Stone-Elander, S. (2012) Site-specifically 11C-labeled Sel-tagged annexin A5 and a size-matched control for dynamic in vivo PET imaging of protein distribution in tissues prior to and after induced cell death. Biochim Biophys Acta 1830, 2562-2573 13.    Wallberg, H., Grafstrom, J., Cheng, Q., Lu, L., Martinsson Ahlzen, H. S., Samen, E., Thorell, J. O., Johansson, K., Dunas, F., Olofsson, M. H., Stone-Elander, S., Arner, E. S., and Stahl, S. (2012) HER2-positive tumors imaged within 1 hour using a site-specifically 11C-labeled Sel-tagged affibody molecule. J Nucl Med 53, 1446-1453 14.    Cheng, Q., Lu, L., Grafstrom, J., Olofsson, M. H., Thorell, J. O., Samen, E., Johansson, K., Ahlzen, H. S., Stone-Elander, S., Linder, S., and Arner, E. S. (2012) Combining [11C]-AnxA5 PET imaging with serum biomarkers for improved detection in live mice of modest cell death in human solid tumor xenografts. PLoS ONE 7, e42151 15.    Arnér, E. S. J. (2009) Focus on mammalian thioredoxin reductases – important selenoproteins with versatile functions. Biochim Biophys Acta 1790, 495-526 16.    Witte, A. B., Anestal, K., Jerremalm, E., Ehrsson, H., and Arner, E. S. (2005) Inhibition of thioredoxin reductase but not of glutathione reductase by the major classes of alkylating and platinum-containing anticancer compounds. Free Radic Biol Med 39, 696-703 17.    Anestål, K., Prast-Nielsen, S., Cenas, N., and Arnér, E. S. J. (2008) Cell Death by SecTRAPs – Thioredoxin Reductase as a Prooxidant Killer of Cells. PLoS ONE 3, e1846 18.    Anestål, K., and Arnér, E. S. J. (2003) Rapid Induction of Cell Death by Selenium-compromised Thioredoxin Reductase 1 but Not by the Fully Active Enzyme Containing Selenocysteine. J Biol Chem 278, 15966-15972 19.    Stafford, W. C., Peng, X., Olofsson, M. H., Zhang, X., Luci, D. K., Lu, L., Cheng, Q., Tresaugues, L., Dexheimer, T. S., Coussens, N. P., Augsten, M., Ahlzen, H.-S. M., Orwar, O., Ostman, A., Stone-Elander, S., Maloney, D. J., Jadhav, A., Simeonov, A., Linder, S., and Arner, E. S. J. (2018) Irreversible inhibition of cytosolic thioredoxin reductase 1 as a mechanistic basis for anticancer therapy. Science translational medicine 10 20.    Ingold, I., Berndt, C., Schmitt, S., Doll, S., Poschmann, G., Buday, K., Roveri, A., Peng, X., Freitas, F. P., Seibt, T., Mehr, L., Aichler, M., Walch, A., Lamp, D., Jastroch, M., Miyamoto, S., Wurst, W., Ursini, F., Amer, E. S. J., Fradejas-Villar, N., Schweizer, U., Zischka, H., Angeli, J. P. F., and Conrad, M. (2018) Selenium Utilization by GPX4 Is Required to Prevent Hydroperoxide-Induced Ferroptosis. Cell 172, 409-+ 21.    Cheng, Q., and Arner, E. S. J. (2017) Selenocysteine Insertion at a Predefined UAG Codon in a Release Factor 1 (RF1)-depleted Escherichia coli Host Strain Bypasses Species Barriers in Recombinant Selenoprotein Translation. Journal of Biological Chemistry 292, 5476-5487    

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