Abstract
We have subjected peas (Pisum sativum L.) to four different oxidative stresses: cold conditions (4 °C) in conjunction with light, treatment with paraquat, fumigation with ozone, and illumination of etiolated seedlings (greening). In crude extracts of leaves from stressed plants, an increase (up to twofold) in activity of glutathione reductase (GR) was observed which was consistent with previous reports from several laboratories. In all cases, except for ozone fumigation, the increase in activity was not due to an elevation in the steady-state levels of GR protein. None of the applied stresses had any effect on steady-state levels of GR mRNA. In contrast to the small increase in GR activity, the K m of GR for glutathione disulphide showed a marked decrease when determined for extracts of stressed leaves, compared with that from unstressed plants. This indicates that GR from stressed plants has an increased affinity for glutathione disulphide. The profile of GR activity bands fractionated on non-denaturing acrylamide gels varied for extracts from differently stressed leaves and when compared with GR from unstressed plants. The changes in GR-band profiles and the alteration in the kinetic properties are best explained as changes in the isoform population of pea GR in response to stress.
Article PDF
Similar content being viewed by others
Avoid common mistakes on your manuscript.
Abbreviations
- GR:
-
glutathione reductase
- GSSG:
-
glutathione disulphide
- Rubisco:
-
Ribulose-1,5-bisphosphate carboxylase-oxygenase
- RNase A/T1:
-
ribonucleases A and T1
References
Accotto, G.P., Donson, J., Mullineaux, P.M. (1989) Mapping of Digitaria streak virus transcripts reveals different RNA species from the same transcription unit. EMBO J. 8, 1033–1039
Alscher, R.G. (1989) Biosynthesis and antioxidant function of glutathione in plants. Physiol. Plant. 77, 457–464
Anderson, J.V., Hess J.L., Chevone, B. (1990) Purification, characterization, and immunological properties for two isoforms of glutathione reductase from eastern white pine. Plant Physiol. 94, 1402–1409
Anderson, J.V., Chevone, B.I., Hess, J.L. (1992) Seasonal variation in the antioxidant system of eastern white pine needles. Plant Physiol. 98, 501–508
Bielawski, V., Joy, K.W. (1986) Properties of glutathione reductase from chloroplasts and roots of pea. Phytochemistry 25, 2261–2265
Bolwell, G.P., Bell, J.N., Cramer, C.L., Schuch, W., Lamb, C.J., Dixon, R.A. (1985) l-phenylalanine ammonia-lyase from Phaseolus vulgaris: Characterisation and differential induction of multiple forms from elicitor-treated cell suspension cultures. Eur. J. Biochem. 149, 411–419
Bradford, M.M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248–254
Cakmak, I., Marschner, H. (1992) Magnesium defficiency and high light intensity enhance activities of superoxide dismutase, ascorbate peroxidase and glutathione reductase in bean leaves. Plant Physiol. 98, 1222–1227
Connell, J.P., Mullet, J.E. (1986) Pea chloroplast glutathione reductase. Purification and characterization. Plant Physiol. 53, 47–51
Creissen, G., Edwards, E.A., Enard, C., Wellburn, A., Mullineaux, P. (1992) Molecular characterization of glutathione reductase cDNAs from pea (Pisum sativum L.) Plant J. 2, 129–131
Dalton, D.A., Russell, S.A., Hanus, F.J., Pascoe, G.A., Evans H.J. (1986) Enzymatic reactions of ascorbate and glutathione that prevent peroxide damage in soybean root nodules. Proc. Natl. Acad. Sci. USA 83, 3811–3815
Dekker, E.L., Woolston, C.J., Xue, Y., Cox, B., Mullineaux, P.M. (1991) Transcript mapping reveals different expression strategies for the bicistronic RNAs of the geminivirus wheat dwarf virus. Nucleic Acids Res 19, 4075–4081
Dixon, R.A., Lamb, C.J. (1990) Molecular communication in interactions between plants and microbial pathogens. Annu. Rev. Plant Physiol. Plant Mol. Biol. 41, 339–367
Dron, M., Clouse, S.D., Dixon, R.A., Lawton, M.A., Lamb, C.J. (1988) Glutathione and fungal elicitor regulation of a plant defense gene promoter in electroporated protoplasts. Proc. Natl. Acad. Sci. USA 85, 6738–6742
Edwards, E.A., Rawsthorne, S., Mullineaux, P.M. (1990) Subcellular distribution of multiple forms of glutathione reductase in leaves of pea (Pisum sativum L.). Planta 180, 278–284
Foster, J.G., Hess, J.L. (1980) Responses of superoxide dismutase and glutathione reductase activities in cotton leaf tissue exposed to an atmosphere enriched in oxygen. Plant Physiol. 66, 482–487
Foster, J.G., Hess, J.L. (1982) Oxygen effects on maize leaf superoxide dismutase and glutathione reductase. Phytochemistry 21, 1527–1532
Foyer, C.H., Halliwell, B. (1976) Presence of glutathione and glutathione reductase in chloroplasts: a proposed role in ascorbic acid metabolism. Planta 133, 21–25
Foyer, C.H., Lelandais, M., Galap, C., Kunert, K.J. (1991) Effects of elevated cytosolic glutathione reductase activity on the cellular glutathione pool and photosynthesis in leaves under normal and stress conditions. Plant Physiol. 97, 863–872
Gebhardt, C., Oliver, J.E., Forde, B.G., Saarelainen, R., Miflin, B.J. (1986) Primary structure and differential expression of glutamine synthetase genes in nodules, roots and leaves of Phaseolus vulgaris. EMBO J. 5, 1429–1435
Gething, M.J., Sambrook, J. (1992) Protein-folding in the cell. Nature 355, 33–45
Grotewald, E., Athama, P., Peterson, T. (1991). Alternatively spliced products of the maize P gene encode proteins with homology to the DNA-binding domain of myb-like transcription factors. Proc. Natl. Acad. Sci. USA 88, 4587–4591
Guy, C.L., Carter, J.V. (1984) Characterisation of partially purified glutathione reductase from cold-hardened and nonhardened spinach leaf tissue. Cryobiology 21, 454–464
Halliwell, B., Foyer, C.H. (1978) Properties and physiological function of a glutathione reductase purified from spinach leaves by affinity chromatography. Planta 139, 9–17
Harker, C.L., Ellis, T.H.N., Coen, E.S. (1990) Identification and genetic regulation of the chalcone synthase multigene family in pea. Plant Cell 2, 185–194
Jahnke, L.S., Hull, M.R., Long, S.P. (1991) Chilling stress and oxygen metabolising enzymes in Zea mays and Zea diploperennis. Plant Cell Environ. 14, 97–104
Kalt-Torres, W., Burke, J.J., Anderson, J.M. (1984) Chloroplast glutathione reductase: Purification and properties. Physiol. Plant. 61, 271–278
Kauffmann, S., Legrand, M., Geoffroy, P., Fritig, B. (1987) Biological function of “pathogenesis-related” proteins: four PR proteins of tobacco have, 1,3-β-glucanase activity. EMBO J. 6, 3209–3212
Klapheck, S., Zimmer, I., Cosse, H. (1990) Scavenging of hydrogen peroxide in the endosperm of Ricinus communis by ascorbate peroxidase. Plant Cell Physiol. 31, 1005–1013
Kuroda, H., Sagisaki, S., Asada, M., Chiba, K. (1991) Peroxidescavenging systems during cold acclimation of apple callus in culture. Plant Cell Physiol. 32, 635–641
Laemmli, U.K. (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227, 680–685
Madamanchi, N.R., Anderson, J.V., Alscher, R.G., Cramer, C.L., Hess, J.L. (1992) Purification of multiple forms of glutathione reductase from pea (Pisum sativum L.) seedlings and enzyme levels in ozone-fumigated pea leaves. Plant Physiol. 100, 138–145
Mahan, J.R., Burke, J.J. (1987) Purification and characterisation of glutathione reductase from corn mesophyll chloroplasts. Physiol. Plant. 71, 352–358
Marana, C., Garcia-Olmedo, F., Carbonero, P. (1990) Differential expression of two types of sucrose synthase-encoding genes in wheat in response to anaerobiosis, cold shock and light. Gene 88, 167–172
Mehlhorn, H., Wellburn, A.R. (1987) Stress ethylene formation determines plant sensitivity to ozone. Nature 327, 417–418
Mehlhorn, H., Cottam, D.A., Lucas, P.W., Wellburn, A.R. (1987) Induction of ascorbate peroxidase and glutathione reductase activities by interactions of mixtures of air pollutants. Free Rad. Res. Commun. 3, 193–197
Melton, D.A., Krieg, P.A., Rebagliati, M.R., Maniatis, T., Zinn, K., Green, M.R. (1984) Efficient in vitro synthesis of biologically active RNA and RNA hybridisation probes from plasmids containing a bacteriophage SP6 promoter. Nucleic Acids Res. 12, 7035–7056
Mullineaux, P.M., Guerineau, F., Accotto, G.P. (1990) Processing of complementary sense RNAs of Digitaria streak virus in its host and in transgenic tobacco. Nucleic Acids Res. 18, 7259–7265
Mullineaux, P.M., Rigden, J.E., Dry, I.B., Krake, L.R., Rezaian, M.A. (1993). Mapping of the polycistronic RNAs of tomato leaf curl geminivirus. Virology 193, 414–423
Nakano, Y., Asada, K. (1980) Spinach chloroplasts scavenge hydrogen peroxide on illumination. Plant Cell Physiol. 21, 1295–1307
Pastori, G.M., Trippi, V.S. (1992) Oxidative stress induces high rate of glutathione reductase synthesis in a drought-resistant maize strain. Plant Cell Physiol. 33, 957–961
Rochester, D.E., Winer, J.A., Shah, D.M. (1986) The structure and expression of maize genes encoding the major heat shock protein, hsp70. EMBO J. 5, 451–458
Schmidt, A., Kunert, K.J. (1987) Antioxidative systems: Defense against oxidative damage in plants. In: Molecular strategies for crop protection, pp. 401–413, Alan R. Liss, New York
Smith, I.K., Vierhaller, T.L., Thorne, C.A. (1988). Assay of glutathione reductase in crude tissue homogenates using 5,5′-dithiobis(2-nitrobenzoic acid). Anal Biochem. 175, 408–413
Sogaard, M., Olsen, F.L., Svensson, B. (1991) C-terminal processing of barley α-amylase I in malt, aleurone protoplasts and yeast. Proc. Natl. Acad. Sci. USA 88, 8140–8144
Tanaka, K., Saji, H., Kondo, N. (1988) Immunological properties of spinach glutathione reductase and inductive biosynthesis of the enzyme with ozone. Plant Cell Physiol. 29, 637–642
Timmerman, K.P. (1989) Molecular characterization of corn glutathione-S-transferase isozymes involved in herbicide detoxification. Physiol. Plant. 77, 465–471
Wingate, V.P.M., Lawton, M.A., Lamb, C.J. (1988) Glutathione elicits a massive and selective induction of plant defense genes. Plant Physiol. 87, 206–210
Zeigler, D.M. (1985) Role of reversible oxidation-reduction of enzyme thiol-disulphides in metabolic regulation. Annu. Rev. Biochem. 54, 305–329
Zinn, K., DiMaio, D., Maniatis, T. (1983). Identification of two distinct regulatory regions adjacent to the human ij-interferon gene. Cell 34, 865–879
Author information
Authors and Affiliations
Additional information
We are grateful to Prof. Alan Wellburn and Dr. Phil Beckett (Division of Biological Sciences, University of Lancaster, UK) for providing ozone-fumigated material and Dr. Jeremy Harbinson for providing material grown at 4° C. This work was supported by a grant-in-aid to the John Innes Institute from the Agricultural and Food Research Council. E.A.E. and C.E. gratefully acknowledge the support of a John Innes Foundation studentship and a European Molecular Biology Organisation Fellowship respectively.
Rights and permissions
About this article
Cite this article
Edwards, E.A., Enard, C., Creissen, G.P. et al. Synthesis and properties of glutathione reductase in stressed peas. Planta 192, 137–143 (1993). https://doi.org/10.1007/BF00198704
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00198704