Abstract
This paper investigates differences in gene expression among the two Thlaspi caerulescens ecotypes La Calamine (LC) and Lellingen (LE) that have been shown to differ in metal tolerance and metal uptake. LC originates from a metalliferous soil and tolerates higher metal concentrations than LE which originates from a non-metalliferous soil. The two ecotypes were treated with different levels of zinc in solution culture, and differences in gene expression were assessed through application of a cDNA microarray consisting of 1,700 root and 2,700 shoot cDNAs. Hybridisation of root and shoot cDNA from the two ecotypes revealed a total of 257 differentially expressed genes. The regulation of selected genes was verified by quantitative reverse transcriptase polymerase chain reaction. Comparison of the expression profiles of the two ecotypes suggests that LC has a higher capacity to cope with reactive oxygen species and to avoid the formation of peroxynitrite. Furthermore, increased transcripts for the genes encoding for water channel proteins could explain the higher Zn tolerance of LC compared to LE. The higher Zn tolerance of LC was reflected by a lower expression of the genes involved in disease and defence mechanisms. The results of this study provide a valuable set of data that may help to improve our understanding of the mechanisms employed by plants to tolerate toxic concentrations of metal in the soil.
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Aharon R, Shahak Y, Wininger S, Bendov R, Kapulnik Y, Galili G (2003) Overexpression of a plasma membrane aquaporin in transgenic tobacco improves plant vigor under favorable growth conditions but not under drought or salt stress. Plant Cell 15:439–447
Alscher RG, Erturk N, Heath LS (2002) Role of superoxide dismutases (SODs) in controlling oxidative stress in plants. J Exp Bot 53:1331–1341
Assunção AGL, Da Costa Martins P, De Folter S, Vooijs R, Schat H, Aarts MGM (2001) Elevated expression of metal transporter genes in three accessions of the metal hyperaccumulator Thlaspi caerulescens. Plant Cell Environ 24:217–226
Assunção AGL, Schat H, Aarts MGM (2003a) Thlaspi caerulescens, an attractive model species to study heavy metal hyperaccumulation in plants. New Phytol 159:351–360
Assunção AGL, Bookum WM, Nelissen HJM, Voojis R, Schat H, Ernst WHO (2003b) Differential metal-specific tolerance and accumulation patterns among Thlaspi caerulescens populations originating from different soil types. New Phytol 159:411–419
Baker AJM (1981) Accumulators and excluders—strategies in the response of plants to heavy metals. J Plant Nutr 3:643–654
Bassi R, Caffarri S (2000) Lhc proteins and the regulation of photosynthetic light harvesting function by xanthophylls. Photosynth Res 64:243–256
Becher M, Talke IN, Krall L, Krämer U (2004) Cross-species microarray transcript profiling reveals high constitutive expression of metal homeostasis genes in shoots of the zinc hyperaccumulator Arabidopsis halleri. Plant J 37:251–268
Belkhadir Y, Subramaniam R, Dangl JL (2004) Plant disease resistance protein signaling: NBS-LRR proteins and their partners. Curr Opin Plant Biol 7:391–399
Benaroya RO, Zamski E, Tel-Or E (2004) L-Myo-inositol 1-phosphate synthase in the aquatic fern Azolla filiculoides. Plant Physiol Biochem 42:97–102
Bones AM, Rossiter JT (1996) The myrosinase–glucosinolate system, its organisation and biochemistry. Physiol Plant 97:194–208
Brown SL, Angle JS, Chaney RL, Baker AJM (1995) Zinc and cadmium uptake by hyperaccumulator Thlaspi caerulescens grown in nutrient solution. Soil Sci Soc Am J 59:125–133
Cakmak I, Braun HJ (2001) Genotypic variation of zinc efficiency. In: Reynolds MP, Ortiz-Monasterio JI, McNab A (eds) Application of physiology in wheat breeding. CIMMYT, Mexico, pp 183–199
Chang YC, Walling LL (1992) Chlorophyll a/b-binding protein genes are differentially expressed during soybean development. Plant Mol Biol 19:217–230
Cunningham SD, Berti WR (1993) Remediation of contaminated soils with green plants: an overview. In Vitro Cell Dev Biol-Plant 29:207–212
Cuypers A, Vangronsveld J, Clijsters H (2001) The redox status of plant cells (AsA and GSH) is sensitive to zinc imposed oxidative stress in roots and primary leaves of Phaseolus vulgaris. Plant Physiol Biochem 39:657–664
Czechowski T, Bari RP, Stitt M, Scheible W-R, Udvardi MK (2004) Real-time RT-PCR profiling over 1400 Arabidopsis transcription factors: unprecedented sensitivity reveals novel root- and shoot-specific genes. Plant J 38:366–379
Dat J, Vandenabeele S, Vranová E, Van Montagu M, Inzé D, Van Breusegem F (2000) Dual action of the active oxygen species during plant stress responses. Cell Mol Life Sci 57:779–795
Dräger DB, Desbrosses-Fonrouge A-G, Krach C, Chardonnes AN, Meyer RC, Saumitou-Laprade P, Krämer U (2004) Two genes encoding Arabidopsis halleri MTP1 metal transport proteins co-segregate with zinc tolerance and account for high MTP1 transcript levels. Plant J 39:425–439
Eckert M, Biela A, Siefritz F, Kaldenhoff R (1999) New aspects of plant aquaporin regulation and specificity. J Exp Bot 50:1541–1545
Edwards R, Dixon DP, Walbot V (2000) Plant glutathione S-transferases: enzymes with multiple functions in sickness and in health. Trends Plant Sci 5:193–198
Engel A, Walz T, Agre P (1994) The aquaporin family of membrane water channels. Curr Opin Struct Biol 4:545–553
Escarré J, Lefèbvre C, Gruber W, Leblanc M, Lepart J, Rivière Y, Delay B (2000) Zinc and cadmium hyperaccumulation by Thlaspi caerulescens from metalliferous and nonmetalliferous sites in the Mediterranean area: implications for phytoextraction. New Phytol 145:429–437
Gelhaye E, Rouhier N, Navrot N, Jacquot JP (2005) The plant thioredoxin system. Cell Mol Life Sci 62:24–35
Gutterson N, Reuber TL (2004) Regulation of disease resistance pathways by AP2/ERF transcription factors. Curr Opin Plant Biol 7:465–471
Hammond JP, Bowen HC, White PJ, Mills V, Pyke KA, Baker AJM, Whiting SN, May ST, Broadley MR (2006) A comparison of the Thaspi caerulescens and Thlaspi arevense shoot transcriptomes. New Phytol 170:239–260
Hardie DG, Carling D, Carlson M (1998) The AMP-activated/SNF1 protein kinase subfamily: metabolic sensors of the eukaryotic cell. Annu Rev Biochem 67:821–855
Hassinen VH, Tervahauta AI, Halimaa P, Plessl M, Peräniemi S, Schat H, Aarts MGM, Servomaa K, Kärenlampi SO (2007) Isolation of Zn-responsive genes from two accessions of the hyperaccumulator plant Thlaspi caerulescens. Planta 225:977–989
Hassinen VH, Tuomainen M, Peräniemi S, Schat H, Kärenlampi SO, Tervahauta AI (2009) Metallothioneins 2 and 3 contribute to the metal-adapted phenotype but are not directly linked to Zn accumulation in the metal hyperaccumulator, Thlaspi caerulescens. J Exp Bot 60:187–196
Herrero J, Al-Shahrour F, Díaz-Uriarte R, Mateos Á, Vaquerizas JM, Santoyo J, Dopazo J (2003) GEPAS: a web-based resource for microarray gene expression data analysis. Nucleic Acids Res 31:3461–3467
Holland MJ (2002) Transcript abundance in yeast varies over six orders of magnitude. J Biol Chem 277:14363–14366
Huang X, von Rad U, Durner J (2002) Nitric oxide induces transcriptional activation of the nitric oxide-tolerant alternative oxidase in Arabidopsis suspension cells. Planta 215:914–923
Inzé D, Van Montagu M (2002) Oxidative stress in plants. Taylor & Francis, London
Jacobs J, Roe JL (2005) SKS6, a multicopper oxidase-like gene, participates in cotyledon vascular patterning during Arabidopsis thaliana development. Planta 222:652–666
Javid-Majd F, Blanchard JS (2000) Mechanistic analysis of the argE-encoded N-acetylornithine deacetylase. Biochemistry 39:1285–1293
Johnson MD, Sussex IM (1995) 1l-Myo-inositol 1-phosphate synthase from Arabidopsis thaliana. Plant Physiol 107:613–619
Kaiser WM, Weiner H, Huber SC (1999) Nitrate reductase in higher plants: a case study for transduction of environmental stimuli into control of catalytic activity. Physiol Plant 105:385–390
Kim D, Gustin JL, Lahner B, Persans MW, Baek D, Yun D-J, Salt DE (2004) The plant CDF family member TgMTP1 from the Ni/Zn hyperaccumulator Thlaspi goesingense acts to enhance efflux of Zn at the plasma membrane when expressed in Saccharomyces cerevisiae. Plant J 39:237–251
Klucas RV, Hanus FJ, Russell SA, Evans HJ (1983) Nickel: a micronutrient element for hydrogen-dependent growth of Rhizobium japonicum and for expression of urease activity in soybean leaves. Proc Natl Acad Sci USA 80:2253–2257
Lee PC, Bochner BR, Ames BN (1983) AppppA, heat-shock stress and cell oxidation. Proc Natl Acad Sci USA 80:7496–7500
Ling H-Q, Koch G, Bäumlein H, Ganal MW (1999) Map-based cloning of chloronerva, a gene involved in iron uptake of higher plants encoding nicotianamine synthase. Proc Natl Acad Sci USA 96:7098–7103
Lingua G, Franchin C, Todeschini V, Castiglione S, Biondi S, Burlando B, Parravicini V, Torrigiani P, Berta G (2008) Arbuscular mycorrhizal fungi differentially affect the response to high zinc concentrations of two registered poplar clones. Environ Pollut 153:137–147
Livak KJ, Schmittgen TD (2001) Analysis of relative gene expression data using real-time quantitative PCR and the \( {2^{ - \Delta \Delta {\text{CT}}}} \) method. Methods 25:402–408
Lombi E, Zhao FJ, Dunham SJ, McGrath SP (2000) Cadmium accumulation in populations of Thlaspi caerulescens and Thlaspi goesingense. New Phytol 145:11–20
Maga JA (1982) Phytate: its chemistry, occurrence, food interactions, nutritional significance, and methods of analysis. J Agricult Food Chem 30:1–9
Mari S, Gendre D, Pianelli K, Ouerdane L, Lobinski R, Briat J-F, Lebrun M, Czernic P (2006) Root-to-shoot long-distance circulation of nicotianamine and nicotianamine-nickel chelates in the metal hyperaccumulator Thlaspi caerulescens. J Exp Bot 57:4111–4122
Marschner H (1995) Mineral nutrition of higher plants, 2nd edn. Academic, London
Meerts P, Van Isacker N (1997) Heavy metal tolerance and accumulation in metallicolous and non-metallicolous populations of Thlaspi caerulescens from continental Europe. Plant Ecol 133:221–231
Milner MJ, Kochian LV (2008) Investigating heavy-metal hpyeraccumulation using Thlaspi caerulescens as a model system. Ann Bot 102:3–13
Moons A (2003) Osgstu3 and osgstu4, encoding tau class glutathione S-transferases, are heavy metal- and hypoxic-stress induced and differentially salt stress-responsive in rice roots. FEBS Lett 553:427–432
Morot-Gaudry-Talarmain Y, Rockel P, Moureaux T, Quilleré I, Leydecker MT, Kaiser WM, Morot-Gaudry JF (2002) Nitrite accumulation and nitric oxide emission in relation to cellular signaling in nitrite reductase antisense tobacco. Planta 215:708–715
Naur P, Petersen BL, Mikkelsen MD, Bak S, Rasmussen H, Olsen CE, Halkier BA (2003) CYP83A1 and CYP83B1, two nonredundant cytochrome P450 enzymes metabolizing oximes in the biosynthesis of glucosinolates in Arabidopsis. Plant Physiol 133:63–72
Nessler CL, Allen RD, Galewsky S (1985) Identification and characterization of latex-specific proteins in opium poppy. Plant Physiol 79:499–504
Nie X, Hill RD (1997) Mitochondrial respiration and hemoglobin gene expression in barley aleurone tissue. Plant Physiol 114:835–840
Noret N, Meerts P, Vanhaelen M, Dos Santos A, Escarré J (2007) Do metal-rich plants deter herbivores? A field test of the defence hypothesis. Oecologia 152:92–100
Ohwaki Y, Kawagishi-Kobayashi M, Wakasa K, Fujihara S, Yoneyama T (2005) Induction of class-1 non-symbiotic hemoglobin genes by nitrate, nitrite and nitric oxide in cultured rice cells. Plant Cell Physiol 46:324–331
Plessl M, Rigola D, Hassinen V, Aarts MGM, Schat H, Ernst D (2005) Transcription profiling of the metal-hyperaccumulator Thlaspi caerulescens (J. & C. PRESL). Z Naturforsch 60c:216–223
Prasad AS (1995) Zinc: an overview. Nutrition 11:93–99
Rask L, Andréasson E, Ekbom B, Eriksson S, Pontoppidan B, Meijer J (2000) Myrosinase: gene family evolution and herbivore defense in Brassicacea. Plant Mol Biol 42:93–113
Reeves RD, Schwartz C, Morel JL, Edmondson J (2001) Distribution and metal-accumulating behaviour of Thlaspi caerulescens and associated metallophytes in France. Int J Phytorem 3:145–172
Rigola D, Fiers M, Vurro E, Aarts MGM (2006) The heavy metal hyperaccumulator Thlaspi caerulescens expresses many species-specific genes, as identified by comparative expressed sequence tag analysis. New Phytol 170:753–766
Rouhier N, Gelhaye E, Gualberto JM, Jordy M-N, De Fay E, Hirasawa M, Duplessis S, Lemaire SD, Frey P, Martin F, Manieri W, Knaff DB, Jacquot J-P (2004) Poplar peroxiredoxin Q. A thioredoxin-linked chloroplast antioxidant functional in pathogen defense. Plant Physiol 134:1027–1038
Sagner S, Kneer R, Wanner G, Cosson J-P, Deus-Neumann B, Zenk MH (1998) Hyperaccumulation, complexation and distribution of nickel in Sebertia acuminata. Phytochemistry 47:339–347
Sakakibara H, Kobayashi K, Deji A, Sugiyama T (1997) Partial characterization of the signaling pathway for the nitrate-dependent expression of genes for nitrogen-assimilatory enzymes using detached maize leaves. Plant Cell Physiol 38:837–843
Schat H, Llugany M, Bernhard R (2000) Metal-specific patterns of tolerance, uptake, and transport of heavy metals in hyperaccumulating and non-hyperaccumulating metallophytes. In: Terry N, Banuelos G (eds) Phytoremediation of contaminated soils and water. CRC, Boca Raton, pp 171–188
Sedbrook JC, Carroll KL, Hung KF, Masson PH, Somerville CR (2002) The Arabidopsis SKU5 gene encodes an extracellular glycosyl phosphatidylinositol-anchored glycoprotein involved in directional root growth. Plant Cell 14:1635–1648
Sharma P, Dubey RS (2005) Modulation of nitrate reductase activity in rice seedlings under aluminium toxicity and water stress: role of osmolytes as enzyme protectant. J Plant Physiol 162:854–864
Sugden C, Donaghy PG, Halford NG, Hardie DG (1999) Two SNF1-related protein kinases from spinach leaf phosphorylate and inactivate 3-hydroxy-3-methylglutaryl-coenzyme A reductase, nitrate reductase, and sucrose phosphate synthase in vitro. Plant Physiol 120:257–274
Suzuki K, Higuchi K, Nakanishi H, Nishizawa NK, Mori S (1999) Cloning of nicotianamine synthase genes from Arabidopsis thaliana. Soil Sci Plant Nutr 45:993–1002
Takahashi M, Terada Y, Nakai I, Nakanishi H, Yoshimura E, Mori S, Nishizawa NK (2003) Role of nicotianamine in the intracellular delivery of metals and plant reproductive development. Plant Cell 15:1263–1280
Tazawa M, Asai K, Iwasaki N (1996) Characteristics of Hg- and Zn-sensitive water channels in the plasma membrane of Chara cells. Bot Acta 109:388–396
Ulmasov T, Ohmiya A, Hagen G, Guilfoyle T (1995) The soybean GH2/4 gene that encodes a glutathione S-transferase has a promoter that is activated by a wide range of chemical agents. Plant Physiol 108:919–927
Unver T, Bozkurt O, Akkaya MS (2008) Identification of differentially expressed transcripts from leaves of the boron tolerant plant Gypsophila perfoliata L. Plant Cell Rep 27:1411–1422
Vallee BL, Falchuk KH (1993) The biochemical basis of zinc physiology. Physiol Rev 73:79–118
van de Mortel JE, Villanueva LA, Schat H, Kwekkeboom J, Coughlan S, Moerland PD, van Themaat EVL, Koorneef M, Aarts MGM (2006) Large expression differences in genes for iron and zinc homeostasis, stress response, and lignin biosynthesis distinguish roots of Arabidopsis thaliana and the related metal hyperaccumulator Thlaspi caerulescens. Plant Physiol 142:1127–1147
Vassilev A, Schwitzguebel J-P, Thewys T, van der Lelie D, Vangronsveld J (2004) The use of plants for remediation of metal-contaminated soils. Sci World J 4:9–34
Wang R, Guegler K, LaBrie ST, Crawford NM (2000) Genomic analysis of a nutrient response in Arabidopsis reveals diverse expression patterns and novel metabolic and potential regulatory genes induced by nitrate. Plant Cell 12:1491–1509
Weber M, Harada E, Vess C, von Roepenack-Lahaye, Clemens S (2004) Comparative microarray analysis of Arabidopsis thaliana and Arabidopsis halleri roots identifies nicotianamine synthase, a ZIP transporter and other genes as potential metal hyperaccumulation factors. Plant J 37:269–281
Weckx JEJ, Clijsters HMM (1997) Zn phytotoxicity induces oxidative stress in primary leaves of Phaseolus vulgaris. Plant Physiol Biochem 35:405–410
Wiebauer K, Ogilvie A, Kersten W (1979) The molecular basis of leucine auxotrophy of quinone-treated Escherichia coli. Active site-directed modification of leucyl-tRNA synthetase by 6-amino-7-chloro-5, 8-dioxoquinoline. J Biol Chem 254:327–332
Zhao FJ, Shen ZG, McGrath SP (1998) Solubility of zinc and interactions between zinc and phosphorus in the hyperaccumulator Thlaspi caerulescens. Plant Cell Environ 21:108–114
Acknowledgements
We thank Ana Assunção for her advice and for her help in providing the Thlaspi plants for our experiments. We would also like to thank the anonymous reviewers for their thoughtful comments and critiques. This work was funded by the European Community (PHYTAC, contract number QLRT-2001-00429).
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Dedicated to Professor Cornelius Lütz on the occasion of his 65th birthday.
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Gene expression ratios of T. caerulescens clones found in the different experiments. The greatest homologues to database are indicated. Italic-typed concentration: induction or repression of a gene; italic-typed ecotype: lower or higher gene expression. Values are average ratios of three biological replicates combined with three complementary swaps. LE, Lellingen; LC, La Calamine; R, hybridisation against root cDNA; S, hybridisation against shoot cDNA; 0, 2, 10, 100, 1,000 indicates the used Zn concentrations (micromolars). Clones 1275–1281 were from A. thaliana. (XLSM 143 kb)
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Plessl, M., Rigola, D., Hassinen, V.H. et al. Comparison of two ecotypes of the metal hyperaccumulator Thlaspi caerulescens (J. & C. PRESL) at the transcriptional level. Protoplasma 239, 81–93 (2010). https://doi.org/10.1007/s00709-009-0085-0
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DOI: https://doi.org/10.1007/s00709-009-0085-0