- Research article
- Open Access
The metal tolerance profile of Thlaspi goesingense is mimicked in Arabidopsis thalianaheterologously expressing serine acetyl-transferase
© Freeman and Salt; licensee BioMed Central Ltd. 2007
- Received: 06 February 2007
- Accepted: 28 November 2007
- Published: 28 November 2007
The Ni hyperaccumulator Thlaspi goesingense is tolerant to Ni ≅ Zn, ≅ Co and slightly resistant to > Cd. We previously observed that elevated glutathione, driven by constitutive activation of serine acetyltransferase (SAT), plays a role in the Ni tolerance of T. goesingense.
Here we show that the elevated shoot concentration of glutathione, previously shown to cause elevated Ni tolerance in Arabidopsis thaliana heterologously expressing T. goesingense mitochondrial serine acetyltransferase (SATm), also causes tolerance to Co and Zn while slightly enhancing resistance to Cd. The level of tolerance afforded to each metal is ranked Ni ≅ Co, > Zn > Cd. The Ni ≅ Co, > Zn tolerances are positively correlated with both the accumulation of glutathione (GSH) and the ability to resist the oxidative damage induced by these different metals. Based on the relative concentrations of each metal used a relatively low level of resistance to Cd was observed in both T. goesingense and TgSATm expressing lines and Cd resistance was least correlated to GSH accumulation.
Such data supports the conclusion that elevated glutathione levels, driven by constitutively enhanced SAT activity in the hyperaccumulator T. goesingense, plays an important role in the Ni, Co and Zn tolerance of this and other Thlaspi species. The hyper-activation of S assimilation through SAT is an excellent strategy for engineering enhanced metal tolerance in transgenic plants potentially used for phytoremediation.
- Photosynthetic Photon Flux
- Metal Tolerance
- Empty Vector Control
- Plant Plate
- Serine Acetyltransferase
The natural phenomenon of heavy metal tolerance in specialized hyperaccumulator plants has sparked the interests of plant physiologists, plant ecologists and evolutionary biologists for over 50 years . The first investigation into metal tolerance present in plant species was by evolutionary ecologists studying primary successionary species that can colonize areas of newly created metaliferous soils such as mine spoils . In recent years, the understanding of the physiological basis of metal(loid) tolerance has increased due to the development of new analytical and molecular technologies [3, 4]. Our research has focused on investigating the physiology of metal tolerance in metal hyperaccumulating plants, which are clearly hyper-tolerant to the metals they hyperaccumulate. However, certain Thlaspi hyperaccumulator species also display co-tolerance to metals not necessarily present in their native habitat . Metal tolerance and metal hyperaccumulation are different but inter-related phenomena, and have been extensively reviewed [1, 4, 6, 7]. In metal hyperaccumulators the genetic determinants of metal tolerance and accumulation segregate independently, confirming that different genes are responsible for these phenomena [8, 9]. Additionally, transcriptional profiling has opened a window into the identity of the genetic changes driving metal hyperaccumulation, by uncovering sets of coordinated gene expression differences that likely control uptake, accumulation and tolerance in hyperaccumulatorsb [10–12]
Metal toxicity in plants is often caused by the production of reactive oxygen species (ROS) [13–17]. Reduced glutathione (GSH) a strong antioxidant can directly reduce some active oxygen species  and also provides the reducing equivalents for the ascorbate-GSH antioxidant cycle . Previous biochemical comparisons between T. goesingense and the closely related non-accumulator A. thaliana, revealed that in T. goesingense constitutively elevated GSH levels, driven by higher activity of the enzyme serine acetyltransferase (SAT), are partly responsible for the elevated Ni tolerance observed in this species . The heterologous expression of T. goesingense SATm in A. thaliana was observed to significantly increase GSH levels, mimicking those found in T. goesingense and consequently conferred increased resistance to Ni . Elevated shoot levels of O-acetyl-L-serine, the product of SAT and a precursor for glutathione biosynthesis has recently been show to also correlate with Ni/Zn hyperaccumulation ability across numerous genera in the Brassicaceae family .
SAT and its product OAS have been shown to control the enzymes responsible for sulfate reduction and cysteine biosynthesis in plants . The production of OAS and cysteine also limit the overall rate of glutathione biosynthesis and the maintenance of an elevated glutathione pool [22, 23]. Sulfur assimilatory mechanisms and the production of GSH are known to be highly induced by both heavy metal treatments and oxidative stress [24, 25]. While oxidative stress tolerances in plants have been increased using transgenic methods to produce elevated levels of OAS, Cys and GSH [26–28], relatively little was known about the effect similar increases might have on the metal tolerance of plants. Increases in these metabolites, through over expression of specific enzymes involved in their biosynthesis, were found to result in Ni tolerance in A. thaliana  and Cd tolerance in B. juncea .
Here we report our observations of the role of GSH in conferring multiple metal tolerances in A. thaliana heterologously expressing TgSATm. These results demonstrate intriguing similarities with the metal tolerance profile observed in the Ni hyperaccumulator T. goesingense with contains naturally enhanced GSH levels. Such data supports our conclusion that constitutively elevated GSH plays an important role in the multi-metal (Ni, Zn and Co), tolerance and Cd resistance observed in T. goesingense and A. thaliana heterologously expressing TgSATm.
Metal tolerance profile of the Ni hyperaccumulator T. goesingense
Metal concentration (μM) which caused 50% inhibition of root growth (I50) for the different metals used on T. goesingense and A. thaliana followed by the relative comparative tolerance index of T. goesingense for different metals.
Comparative Tolerance Index (I50 Thlaspi/I50 Arabidopsis)
Ni > 6.28
Zn > 5.09
Metal tolerance profile of A. thaliana heterologously expressing T. goesingenseserine acetyl-transferase
Shoot material from the TgSATm lines and empty vector controls grown at 20 mm and 35 mm from the disc were harvested for ICPMS analysis to investigate the respective metal concentrations. Interestingly, although tolerance to Ni, Co and Zn was increased in lines heterologously expressing TgSAT m , these tolerant lines had little or no increase in shoot accumulation of Ni, Co or Cd. The only exception to this was for Zn, because both the highest (S 4–9), and intermediate (S 3-1) SAT expressing and GSH accumulating TgSATm lines harvested from the 20 mm ring had a 1.3–1.5 fold higher shoot Zn concentration (8,936 ± 760 & 10,050 ± 458 μg Zn g-1 Dwt respectively) compared to the low (S 5-4) and non-expressing line (E 1–5) at (6,704 ± 656 & 6,644 ± 94 μg Zn g-1 Dwt respectively). While the tolerant SAT lines did not accumulate more metal than the others except for Zn the overall levels were all quite high, for Ni (~1,200), Zn (~6,000–10,000), Co (~2,000) and Cd (~2,000) μg g-1 Dwt.
Association of GSH with tolerance to Ni, Co, Zn and enhanced resistance to Cd
The T. goesingense mitochondrial version of SAT was chosen for use in this study over that of the chloroplast and the cytosolic versions because it conveyed a high level of Ni tolerance to A. thaliana , and because of its high sequence similarity (95.0% identity) to the A. thaliana mitochondrial homolog At3g13110 . Additionally, in E. coli, TgSATm conferred enhanced resistance to Ni , which was originally used to clone the three different SAT cDNAs from T. goesingense . Furthermore, overexpression of the cytoplasmic, mitochondrial, or chloroplastic SAT from T. goesingense or the chloroplastic SAT from A. thaliana in E. coli all conferred approximately equal tolerance to Ni and Co . The mitochondrial isoform of SAT (TgSAT-m) was also chosen for overexpression in this study because overexpression of the Cys-insensitive A. thaliana homolog AtSAT-m (SAT A) had been shown to achieve significant elevations in Cys and GSH concentrations, when overexpressed in tobacco and targeted to either the plastid or the cytosol . In addition to making more OAS, Cys and GSH in the mitochondria, which can then be exported into the cytosol and other organelles, over-expression of TgSATm in the mitochondria could lower Ni induced ROS formation, so the electron transport chain can still provide cells with the energy needed to function.
The hyperaccumulator T. goesingense has significant tolerance to the growth inhibitory effects of elevated Ni, Zn and Co, with a slight resistance to Cd (Fig. 1 and Table 1), and in part this appears due to an enhanced ability to resist the oxidizing effects of Ni, Zn and Co (Fig 2). The elevated levels of GSH in T. goesingense are thought to contribute to the Ni tolerance of this plant, via enhancing resistance to the oxidizing effects of this metal . We hypothesize that this elevated GSH also causes the elevated tolerance to Co and Zn, but only low level resistance to Cd. To address this question we utilized transgenic A. thaliana heterologously expressing TgSAT m that had previously been characterized to contain elevated shoot concentrations of GSH that confer enhanced resistance to Ni .
In addition to Ni our analysis of the metal tolerance of these same plants showed them to also have tolerance to growth in the presence of elevated Co and Zn while causing a low level of resistance to Cd, when compared to the control lines (Fig. 3 &4). Interestingly, the high and intermediate TgSATm expressing lines were found to accumulate ~1.4 fold more Zn when compared to the low and non-expressing lines. This suggests that increasing GSH and thus Zn tolerance might positively influence the Zn accumulation mechanisms in plants.
Furthermore, these transgenic plants also showed increased resistance to the lipid peroxidizing effects of Co and Zn (Fig. 5), suggesting that at least one of the roles of GSH in this enhanced Co and Zn resistance is as an antioxidant. These results mirror those previously observed for Ni . However, the elevated GSH concentration in these lines had no effect on levels of lipid peroxidation in plants grown in the presence of Cd, suggesting that GSH is conferring enhanced tolerance to Cd via an unknown alternative pathway potentially through an increased ability to synthesize phytochelatins (PC). Cd resistance was quite low considering that both T. goesingense and the TgSATm could tolerate growth on plates containing at least 4 times higher concentrations of Ni, Co and Zn compared to Cd.
Our results establish that heterologous expression of TgSAT m also enhances tolerance to other oxidative stresses caused by hydrogen peroxide and high light treatment (Fig. 6). The hyperaccumulator T. goesingense, which is known to have constitutively elevated SAT activity and accumulate, elevated GSH , also shows high tolerance to hydrogen peroxide treatment (Fig. 6).
Analysis of the ability of GSH to confer metal tolerance in these transgenic lines also revealed that in shoots GSH is most effective at conferring tolerance to Ni = Co > Zn > Cd (Fig. 7A). This activity closely follows the levels of lipid peroxidation produced by growth on these three metals (Fig. 5). GSH production was least effective at conferring tolerance to Cd (Fig. 7A), which was exemplified by the no reduction in lipid peroxidation after Cd treatment (Fig. 5). In roots GSH conferred tolerance to Co > Ni = Zn > Cd (Fig. 7B).
In both T. goesingense and the TgSATm transgenics, SAT enhanced Ni, resistance appears not to involve coordination by PC. This is fully supported by a complete lack of thiol coordination to Ni in these plants [19, 31] and also in E. coli heterologously expressing the same gene . Additionally, Zn is also not coordinated by PC or thiols in the Zn tolerant hyperaccumulator T. caerulescens . It has also been determined genetically in the A. thaliana PCS null mutant cad1, which has no measurable PC, that PC are not involved in either Ni or Zn resistance in A. thaliana but are directly needed for Cd resistance [33–36]. While PC are clearly not involved in resistance to Ni and Zn in A. thaliana, Co also failed to activate A. thaliana PC synthesis in assays of purified AtPCS1 activity . Therefore Given the chemical similarity between Ni and Co we can speculate that the enhanced Co resistance observed in the transgenic plants is due to the direct antioxidant effect of increased GSH.
Our findings establish a strong connection between elevated shoot GSH and enhanced tolerance to the lipid peroxidizing effects of Ni = Co > Zn in both T. goesingense and transgenic A. thaliana containing elevated GSH concentrations. Such a link is supported by the observation that elevated GSH levels in both transgenic A. thaliana and T. goesingense correlate with enhanced resistance to the peroxidizing effects caused by hydrogen peroxide and high light treatments.
We therefore conclude that the multiple metal tolerances to Ni, Co and Zn observed in T. goesingense are due to the constitutively elevated shoot GSH levels observed in this hyperaccumulator. These multiple metal tolerances are mimicked in Arabidopsis through heterologous expression of the TgSATm gene which results in elevated GSH accumulation in leaves and enhanced oxidative stress resistance.
Plant plate growth
Plants were germinated and grown on plates containing 1/2 strength MS + Gamborgs B5 medium in 0.8% plant tissue grade agar alone or with the different metals under 16 hr white light (120 μmol m-2 s-1 of photosynthetic photon flux (PPF)). ICP-MS analysis of the different metal levels was performed on all dried shoot tissue as previously described (Lahner et al., 2003).
Metal tolerance assays
Metal resistance in A. thaliana was tested on agar plates (100 × 25 mm) containing 1/2 strength MS + Gamborgs B5 medium in 0.8% plant tissue grade agar. Seeds from A. thaliana heterologously expressing T. goesingense SATm and empty vector control lines were spotted in concentric rings radiating from the center of the plate where a filter paper disc was placed. Onto the disc 100 μL of 100 mM Ni(NO3)2, 100 mM Co (NO3)2, 400 mM Zn (NO3)2 or 50 mM Cd (NO3)2 was pipetted and the plates incubated at 24°C/20°C, 10 h/14 h light/dark, 120 μmol m-2 s-1 PPF. After 13 days growth from imbibition the percentage of upright seedlings with fully expanded cotyledons, secondary leaves and hooked roots were recorded for each ring along with the distance in mm from the metal soaked filter disc. Seedlings from both 20 and 35 mm rings were then harvested for ICP-MS analysis of metal levels. Metal analysis was performed on dried shoot tissue as previously described .
Hydrogen peroxide treatments
Hydrogen peroxide treatments involved cutting 1 cm diameter leaf discs from the center of intact mature leaves. Discs were floated on either H2O2 or water for 24 h at 24°C in constant light (180 μmol m-2 s-1 PPF). Percent total chlorophyll was measured at 654 nm following extraction in 96% ethanol , and normalized to chlorophyll observed in the water treated tissue. The chlorophyll contents for the control (H2O) treatments were approximately the same per gram fresh weight for all plants used. Total Chlorophyll mg g-1 fwt.; T. goesingense = 1.55 ± 0.32, A. thaliana = 1.57 ± 0.13, Transgenic TgSATm = 1.58 ± 0.13.
Quantification of total lipid peroxidation
Peroxidized lipids were assayed as the presence of thiobarbituric acid reactive species (TBARS). Plants were grown for 17 days under 16 hr white light (120 μmol m-2 s-1 PPF) on vertical agar plates containing 1/2 strength MS + Gamborgs B5 medium in 0.8% plant tissue grade agar. After 4 or 8 days transfer to 1/2 strength MS + Gamborgs B5 medium in 0.8% plant tissue grade agar plates, containing either Ni, Co, Zn or Cd, shoot tissue was harvested and assayed for TBARS as described previously . During the period of metal exposure all plants remained viable and continued growing
Root length inhibition
T. goesingense and A. thaliana plants were germinated and grown on vertical plates containing 1/2 strength MS + Gamborgs B5 medium in 0.8% agar with various concentrations of Ni, Co, Zn and Cd for 18 days under 16 hr white light (120 μmol m-2 s-1 PPF). The (I50) values were calculated by taking the metal concentrations in plates which caused 50% inhibition of root growth for the different metals used on both T. goesingense and A. thaliana. The maximum normal root growth was measured on the control plates for this experiment in the absence of any metal.
1 This work was supported by the U.S. National Science Foundation (grant nos. 0196310-IBN and 0129747-IBN to D.E.S.) and by the Indiana 21st Century Research and Technology Fund. Special thanks to the Purdue Ionomics Center, especially Brett Lahner for inductively coupled plasma analyses.
- Baker AJM: Metal Tolerance. New Phytol. 1987, 106: 93-111.View ArticleGoogle Scholar
- Bradshaw AD, McNeilly T, Putwain PD: The essential qualities. Heavy metal tolerance in plants: evolutionary aspects. Edited by: Shaw AJ. CRC Press Inc. Boca Raton, 1989:324-334.Google Scholar
- Clemens S: Molecular mechanisms of plant metal tolerance and homeostasis. Planta. 2001, 212: 475-486. 10.1007/s004250000458.PubMedView ArticleGoogle Scholar
- Meharg AA: Mechanisms of plant resistance to metal and metalloid ions and potential biotechnological applications. Plant and Soil. 2004, 0: 1-11.Google Scholar
- Baker AJM, Reeves RD, Hajar SM: Heavy metal accumulation and tolerance in British populations of the metallophyte Thlaspi caerulescens J. & C. Presl (Brassicaceae). New Phytol. 1994, 127: 6l-68. 10.1111/j.1469-8137.1994.tb04259.x.View ArticleGoogle Scholar
- Persans MW, Salt DE: Possible molecular mechanisms involved in nickel, zinc and selenium hyperaccumulation in plants. Biotechnol Genet Eng Rev. 2000, 17: 389-413.PubMedView ArticleGoogle Scholar
- Salt DE, Kramer U: Mechanisms of Metal Hyperaccumulation in Plants. Phytoremediation of Toxic Metals. Edited by: Raskin I, Ensley BD. John Wiley and Sons Inc, 2000:231-246.Google Scholar
- Macnair MR, Bert V, Huitson SB, Saumitou-Laprade P, Petit D: Zinc tolerance and hyperaccumulation are genetically independent characters. Proc R Soc Lond Biol Sci. 1999, 266: 2175-2179. 10.1098/rspb.1999.0905.View ArticleGoogle Scholar
- Assuncao AGL, Ten Bookum WM, Nelissen JM, Vooijs R, Schat H, Ernst WHO: A cosegregation analysis of zinc (Zn) accumulation and Zn tolerance in the Zn hyperaccumulator Thlaspi caerulescens. New Phytol. 2003, 159: 45-56.Google Scholar
- Becher M, Talke IN, Krall L, Krämer U: Cross-species microarray transcript profiling reveals high constituative expression of metal homeostasis genes in shoots of the zinc hyperaccumulator Arabidopsis halleri. Plant J. 2004, 37: 251-268.PubMedView ArticleGoogle Scholar
- Weber M, Harada E, Vess C, Roepenack-Lahaye EV, Clemens S: 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. 2004, 37: 269-281. 10.1111/j.1365-313X.2003.02013.x.PubMedView ArticleGoogle Scholar
- Hammond JP, Bowen HC, White PJ, Mills V, Pyke KA, Baker AJM, Whiting SN, May ST, Broadley A, Martin R: A comparison of the Thlaspi caerulescens and Thlaspi arvense shoot transcriptomes. New Phytol. 2006, 170: 239-260. 10.1111/j.1469-8137.2006.01662.x.PubMedView ArticleGoogle Scholar
- Baccouch S, Chaoui A, El Ferjani E: Nickel-induced oxidative damage and antioxidant responses in Zea mays shoots. Plant Physiol Biochem. 1998, 36: 689-694. 10.1016/S0981-9428(98)80018-1.View ArticleGoogle Scholar
- Briat JF, Lebrun M: Plant responses to metal toxicity. C R Acad Sci III. 1999, 322: 43-54.PubMedView ArticleGoogle Scholar
- Dat J, Vandenabeele S, Vranova E, Van Montagu M, Inze D, Van Breusegem F: Review, Dual action of the active oxygen species during plant stress responses. Cell Mol Life Sci. 2000, 57: 779-795. 10.1007/s000180050041.PubMedView ArticleGoogle Scholar
- Noctor G, Foyer CH: Ascorbate and Glutathione: Keeping Active Oxygen Under Control. Annu Rev Plant Physiol Plant Mol Biol. 1998, 49: 249-279. 10.1146/annurev.arplant.49.1.249.PubMedView ArticleGoogle Scholar
- Schutzendubel A, Polle A: Plant responses to abiotic stresses: heavy metal-induced oxidative stress and protection by mycorrhization. J Exp Bot. 2002, 53: 1351-1365. 10.1093/jexbot/53.372.1351.PubMedView ArticleGoogle Scholar
- Noctor G, Strohm M, Jouanin L, Kunert KJ, Foyer CH, Rennenberg H: Synthesis of Glutathione in Leaves of Transgenic Poplar Overexpressing [gamma]-Glutamylcysteine Synthetase. Plant Physiol. 1996, 112: 1071-1078.PubMedPubMed CentralGoogle Scholar
- Freeman JL, Persans MW, Nieman K, Albrecht C, Peer W, Pickering IJ, Salt DE: Increased glutathione biosynthesis plays a role in nickel tolerance in Thlaspi nickel hyperaccumulators. Plant Cell. 2004, 16: 2176-2191. 10.1105/tpc.104.023036.PubMedPubMed CentralView ArticleGoogle Scholar
- Peer WA, Mahmoudian M, Freeman JL, Lahner B, Richards EL, Reeves RD, Murphy AS, Salt DE: Assessment of Plants from the Brassicaceae Family as Genetic Models for the Study of Nickel and Zinc Hyperaccumulation. New Phytologist. 2006, 172: 248-260. 10.1111/j.1469-8137.2006.01820.x.PubMedView ArticleGoogle Scholar
- Wirtz M, Hell R: Production of cysteine for bacterial and plant biotechnology application of cysteine feedback-insensitive isoforms of serine acetyltransferase. Amino Acids. 2003, 24: 195-203.PubMedGoogle Scholar
- Barroso C, Vega JM, Gotor C: A new member of the cytosolic O-acetylserine(thiol)lyase gene family in Arabidopsis thaliana. FEBS Lett. 1995, 363: 1-5. 10.1016/0014-5793(95)00255-8.PubMedView ArticleGoogle Scholar
- Meyer AJ, Fricker MD: Control of demand-driven biosynthesis of glutathione in green Arabidopsis suspension culture cells. Plant Physiol. 2002, 130: 1927-1937. 10.1104/pp.008243.PubMedPubMed CentralView ArticleGoogle Scholar
- Xiang C, Oliver DJ: Glutathione metabolic genes coordinately respond to heavy metals and jasmonic acid in Arabidopsis. Plant Cell. 1998, 10: 1539-1550. 10.1105/tpc.10.9.1539.PubMedPubMed CentralView ArticleGoogle Scholar
- Howarth JR, Dominguez-Solis JR, Gutierrez-Alcala G, Wray JL, Romero LC, Gotor C: The serine acetyltransferase gene family in Arabidopsis thaliana and the regulation of its expression by cadmium. Plant Mol Biol. 2003, 51: 589-598. 10.1023/A:1022349623951.PubMedView ArticleGoogle Scholar
- Blasczyk A, Brodzik R, Sirko A: Increased resistance to oxidative stress in transgenic tobacco plants over expressing bacterial serine acetyltransferase. Plant J. 1999, 20: 237-243. 10.1046/j.1365-313x.1999.00596.x.View ArticleGoogle Scholar
- Foyer CH, Souriau N, Perret S, Lelandais M, Kunert KJ, Pruvost C, Jouanin L: Overexpression of glutathione reductase but not glutathione synthetase leads to increases in antioxidant capacity and resistance to photoinhibition in poplar trees. Plant Physiol. 1995, 109: 1047-1057. 10.1104/pp.109.3.1047.PubMedPubMed CentralView ArticleGoogle Scholar
- Youssefian S, Nakamura M, Orudgev E, Kondo N: Increased cysteine biosynthesis capacity of transgenic tobacco overexpressing an O-acetylserine(thiol) lyase modifies plant responses to oxidative stress. Plant Physiol. 2001, 126: 1001-1011. 10.1104/pp.126.3.1001.PubMedPubMed CentralView ArticleGoogle Scholar
- Zhu YL, Pilon-Smits EA, Tarun AS, Weber SU, Jouanin L, Terry N: Cadmium tolerance and accumulation in Indian mustard is enhanced by overexpressing gamma-glutamylcysteine synthetase. Plant Physiol. 1999, 121: 1169-1178. 10.1104/pp.121.4.1169.PubMedPubMed CentralView ArticleGoogle Scholar
- Freeman JL, Persans MW, Nieman K, Salt DE: Nickel and Cobalt Resistance Engineered in Escherichia coli by Overexpression of Serine Acetyltransferase from the Nickel Hyperaccumulator Plant Thlaspi goesingense. Applied and Environmental Microbiology. 2005, 71: 8627-8633. 10.1128/AEM.71.12.8627-8633.2005.PubMedPubMed CentralView ArticleGoogle Scholar
- Kramer U, Pickering IJ, Prince RC, Raskin I, Salt DE: Subcellular localization and speciation of nickel in hyperaccumulator and non-accumulator Thlaspi species. Plant Physiol. 2000, 122: 1343-1353. 10.1104/pp.122.4.1343.PubMedPubMed CentralView ArticleGoogle Scholar
- Salt DE, Prince RC, Baker AJM, Raskin I, Pickering IJ: Zinc ligands in the metal hyperaccumulator Thlaspi caerulescens as determined using X-ray absorption spectroscopy. Env Sci Tech. 1999, 33: 713-717. 10.1021/es980825x.View ArticleGoogle Scholar
- Howden R, Cobbett CS: Cadmium-Sensitive Mutants of Arabidopsis thaliana. Plant Physiol. 1992, 100 (1): 100-107.PubMedPubMed CentralView ArticleGoogle Scholar
- Howden R, Goldsbrough PB, Andersen CR, Cobbett CS: Cadmium-sensitive, cad1 mutants of Arabidopsis thaliana are phytochelatin deficient. Plant Physiol. 1995, 107: 1059-1066. 10.1104/pp.107.4.1059.PubMedPubMed CentralView ArticleGoogle Scholar
- Ha S-B, Smith AP, Howden R, Dietrich WM, Bugg S, O'Connell MJ, Goldsbrough PB, Cobbett CS: Phytochelatin synthase genes from Arabidopsis and the yeast Schizosaccharomyces pombe. Plant Cell. 1999, 11: 1153-1163. 10.1105/tpc.11.6.1153.PubMedPubMed CentralView ArticleGoogle Scholar
- Cobbett CS: Phytochelatins and their role in heavy metal detoxification. Plant Physiol. 2000, 123: 825-832. 10.1104/pp.123.3.825.PubMedPubMed CentralView ArticleGoogle Scholar
- Vatamaniuk OK, Mari S, Lu Y-P, Rea PA: Mechanism of heavy metal ion activation of phytochelatin (PC) synthase. J Biol Chem. 2000, 275: 31451-31459. 10.1074/jbc.M002997200.PubMedView ArticleGoogle Scholar
- Lahner B, Gong J, Mahmoudian M, Smith EL, Abid KB, Rogers EE, Guerinot ML, Harper JF, Ward JM, McIntyre L, Schroeder JI, Salt DE: Genomic scale profiling of nutrient and trace elements in Arabidopsis thaliana. Nat Biotechnol. 2003, 21: 1215-1221. 10.1038/nbt865.PubMedView ArticleGoogle Scholar
- Wintermans JFGM, De Mots A: Spectrophotometric Characteristics of Chlorophylls a and b and their Pheophytins in Ethanol. Biochim Biophys Acta. 1965, 109: 448-453. 10.1016/0926-6585(65)90170-6.PubMedView ArticleGoogle Scholar
- Murphy AS, Eisinger WR, Shaff JE, Kochian LV, Taiz L: Early copper-induced leakage of K+ from Arabidopsis seedlings is mediated by ion channels and coupled to citrate efflux. Plant Physiol. 1999, 121: 1375-1382. 10.1104/pp.121.4.1375.PubMedPubMed CentralView ArticleGoogle Scholar
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.