Structural and functional characterization of tree proteins involved in redox regulation: a new frontier in forest science

Dec 2014

Key message This paper describes how the combination of genomics, genetic engineering, and 3D structural characterization has helped clarify the redox regulatory networks in poplar with consequences not only in system biology in plants but also in bacteria and mammalian systems. Context Tree genomes are increasingly available with a large number of orphan genes coding for proteins, the function of which is still unknown. Aims and methods Modern techniques of genome analysis coupled with recombinant protein technology and massive 3D structural determination of tree proteins should help elucidate the function of many of the proteins encoded by orphan genes. X-ray crystallography and NMR will be the methods of choice for protein structure determination. Results In this review, we provide examples illustrating how the above-mentioned techniques improved our understanding of redox regulatory circuits in poplar, the first forest tree species sequenced. We showed that poplar peroxiredoxins use either thioredoxin or glutaredoxin as electron donors to reduce hydrogen peroxide. That glutaredoxin could be a reductant was unknown at the time of this discovery even in other biological organisms and was later confirmed notably by the observation that the two genes are fused in some bacteria and by the resolution of the structure of the bacterial hybrid protein. Similarly, genome analysis coupled to in vitro analysis of enzymatic properties led to the discovery that some plant methionine sulfoxide reductases can also use both thioredoxins and glutaredoxins as electron donors. Besides their disulfide reductase activity, it has been demonstrated that some poplar glutaredoxins are also involved in iron-sulfur center biogenesis and assembly. The original 3D structure determination has been made with poplar glutaredoxin C1 and then confirmed in a variety of other biological organisms including human. Our work also showed that in plants, so-called glutathione peroxidases use thioredoxins and not glutathione as electron donors. This is true for all non-selenocysteine-containing glutathione peroxidases. Finally, connections between the thioredoxin and glutaredoxin systems have been elucidated through the study of atypical poplar thioredoxins. Conclusion Altogether, these data illustrate how the combination of genetic engineering and structural biology improves our understanding of biological processes and helps fuel systems biology for trees and other biological species.

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Structural and functional characterization of tree proteins involved in redox regulation: a new frontier in forest science

Annals of Forest Science (2016) 73:119–134 DOI 10.1007/s13595-014-0442-9 REVIEW PAPER Structural and functional characterization of tree proteins involved in redox regulation: a new frontier in forest science Jean-Pierre Jacquot & Jérémy Couturier & Claude Didierjean & Eric Gelhaye & Mélanie Morel-Rouhier & Arnaud Hecker & Christophe Plomion & Desirée D. Gütle & Nicolas Rouhier Received: 12 September 2014 / Accepted: 14 November 2014 / Published online: 5 December 2014 # INRA and Springer-Verlag France 2014 Abstract & Key message This paper describes how the combination of genomics, genetic engineering, and 3D structural characterization has helped clarify the redox regulatory networks in poplar with consequences not only in system biology in plants but also in bacteria and mammalian systems. & Context Tree genomes are increasingly available with a large number of orphan genes coding for proteins, the function of which is still unknown. & Aims and methods Modern techniques of genome analysis coupled with recombinant protein technology and massive 3D structural determination of tree proteins should help elucidate the function of many of the proteins encoded by orphan genes. X-ray crystallography and NMR will be the methods of choice for protein structure determination. & Results In this review, we provide examples illustrating how the above-mentioned techniques improved our understanding of redox regulatory circuits in poplar, the first forest tree species sequenced. We showed that poplar peroxiredoxins use either thioredoxin or glutaredoxin as electron donors to Handling Editor: Jean-Michel Leban Executive summary This review describes the successful use of technologies to overproduce and purify recombinant enzymes from trees with the aim of solving their 3D structures and identifying their molecular interactions either with other proteins or with potential substrates. Currently only ca a dozen of tree genomes have been annotated and released including six forest species, but this will change rapidly with the oncoming release of the chestnut and oak genomes notably. The availability of additional genomes will open the way to identifying the function of a large number of orphan gene products, one of the ways to characterize these functions being to produce and analyze the corresponding protein products. J.<P. Jacquot (*) : J. Couturier : E. Gelhaye : M. Morel-Rouhier : A. Hecker : D. D. Gütle : N. Rouhier Université de Lorraine, Interactions Arbres - Microorganismes, UMR1136, 54500 Vandoeuvre-lès-Nancy, France e-mail: J.<P. Jacquot : J. Couturier : E. Gelhaye : M. Morel-Rouhier : A. Hecker : D. D. Gütle : N. Rouhier INRA, Interactions Arbres - Microorganismes, UMR1136, 54280 Champenoux, France C. Didierjean Université de Lorraine, CRM2, Equipe BioMod, UMR 7036, Faculté des Sciences et Technologies, BP 70239, 54506 Vandoeuvre-lès-Nancy, France C. Didierjean CNRS, CRM2, Equipe BioMod, UMR 7036, Faculté des Sciences et Technologies, BP 70239, 54506 Vandoeuvre-lès-Nancy, France C. Plomion INRA, UMR1202, BIOGECO, 33610 Cestas, France D. D. Gütle Plant Biotechnology, Faculty of Biology, Universität Freiburg, Schänzlestrasse 1, 79104 Freiburg, Germany D. D. Gütle Spemann Graduate School of Biology and Medicine (SGBM), University of Freiburg, Albertstr. 19A, 79104 Freiburg, Germany 120 reduce hydrogen peroxide. That glutaredoxin could be a reductant was unknown at the time of this discovery even in other biological organisms and was later confirmed notably by the observation that the two genes are fused in some bacteria and by the resolution of the structure of the bacterial hybrid protein. Similarly, genome analysis coupled to in vitro analysis of enzymatic properties led to the discovery that some plant methionine sulfoxide reductases can also use both thioredoxins and glutaredoxins as electron donors. Besides their disulfide reductase activity, it has been demonstrated that some poplar glutaredoxins are also involved in iron-sulfur center biogenesis and assembly. The original 3D structure determination has been made with poplar glutaredoxin C1 and then confirmed in a variety of other biological organisms including human. Our work also showed that in plants, socalled glutathione peroxidases use thioredoxins and not glutathione as electron donors. This is true for all nonselenocysteine-containing glutathione peroxidases. Finally, connections between the thioredoxin and glutaredoxin systems have been elucidated through the study of atypical poplar thioredoxins. & Conclusion Altogether, these data illustrate how the combination of genetic engineering and structural biology improves our understanding of biological processes and helps fuel systems biology for trees and other biological species. Keywords 3D protein structure . Genome sequence . Glutaredoxin . Redox . Thioredoxin . Poplar 1 Introduction There are currently more than 50 plant genomes sequenced and published (Michael and Jackson 2013; see also: http://en. wikipedia.org/wiki/List_of_sequenced_plant_genomes), and of these, more than ten concern tree species. The majority of those are constituted by fruit trees (peach, plum, pear, sweet orange, clementine, apple, coffee, papaya, and cocoa tree). The first forest tree genome published is Populus trichocarpa (Tuskan et al. 2006) followed by Picea abies (Nystedt et al. 2013), Hevea brasiliensis (Rahman et al. 2013), Picea glauca (Birol et al. 2013), Pinus taeda (Neale et al. 2014), and Eucalyptus grandis (Myburg et al. 2014). The analysis of the first tree genomes has helped in identifying the total number of genes present in these genomes and also a number of orphan genes that have no counterpart in other biological organisms. A recent estimate of the number of orphan genes in poplar is 44 %, a value much higher than the 14 % estimation in Arabidopsis (Guo et al. 2007; Feldmann and Goff 2013). The mechanisms leading to orphan gene emergence have been discussed in a number of biological systems (Wissler et al. 2013). In order to understand the functions and interaction properties of the proteins encoded J.-P. Jacquot et al. by these genes (if any), one possibility is to produce the corresponding recombinant proteins. The elucidation of their 3D structures together with molecular docking is one possibility that can be used to reach that understanding. Other techniques as the yeast two-hybrid system screen and coimmunoprecipitation can also help in identifying the potential ligands either metabolites or macromolecules as other proteins, carbohydrates, or lipids. Together with the results of transcriptomic and proteomic experiments, these approaches will help decipher metabolic and regulation networks. We illustrate here the use of genetic engineering and 3D structure determination to describe the redox interaction networks in trees, concentrating on the glutathione/glutaredoxin- (Grx) and thioredoxin- (Trx) dependent pathways in poplar. Most of the enzymes pre (...truncated)


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Jean-Pierre Jacquot, Jérémy Couturier, Claude Didierjean, Eric Gelhaye, Mélanie Morel-Rouhier, Arnaud Hecker, Christophe Plomion, Desirée D. Gütle, Nicolas Rouhier. Structural and functional characterization of tree proteins involved in redox regulation: a new frontier in forest science, 2016, pp. 119-134, Volume 73, Issue 1, DOI: 10.1007/s13595-014-0442-9