Derivatization of Phytochelatins from Silene vulgaris, Induced upon Exposure to Arsenate and Cadmium:  Comparison of Derivatization with Ellman's Reagent and Monobromobimane

Sneller, F. E. C.; Heerwaarden, L. M. Van; Koevoets, Paul L. M.; Vooijs, Riet; Schat, Henk; Verkleij, J. A. C.

doi:10.1021/jf9903105

Cited by 173 publications

(142 citation statements)

References 27 publications

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“…Thiol-containing compounds were extracted following the method of Sneller et al (2000) with some modifications. Frozen (-80ºC) root and shoot samples were ground in liquid nitrogen (N 2 ) using a mortar and pestle, and 0.10 g of each sample was immediately placed in an individual microcentrifuge tube containing 1.5 mL of 6.3 mM DTPA with 0.1% (v/v) TFA and 25 µL of 20 mM TCEP (4ºC).…”

Section: Extraction Of Thiol-containing Moleculesmentioning

confidence: 99%

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Reduced translocation of cadmium from roots is associated with increased production of phytochelatins and their precursors

Akhter

McGarvey

Macfie

2012

Journal of Plant Physiology

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SummaryCadmium (Cd) is a non-essential trace element and its environmental concentrations are approaching toxic levels, especially in some agricultural soils. Understanding how and where Cd is stored in plants is important for ensuring food safety. In this study, we examined two plant species that differ in the distribution of Cd among roots and leaves. Lettuce and barley were grown in nutrient solution under two conditions: chronic (4 week) exposure to a low, environmentally relevant concentration (1.0 µM) of Cd and acute (1 h) exposure to a high concentration (5.0 mM) of Cd. Seedlings grown in solution containing 1.0 µM CdCl 2 did not show symptoms of toxicity and, at this concentration, 77% of the total Cd was translocated to leaves of lettuce, whereas only 24% of the total Cd was translocated to barley leaves. We tested the hypothesis that differential accumulation of Cd in roots and leaves is related to differential concentrations of phytochelatins (PCs), and its precursor peptides. The amounts of PCs and their precursor peptides in the roots and shoots were measured using HPLC. Each of PC 2-4 was synthesized in the barley root upon chronic exposure to Cd and did not increase further upon acute exposure. In the case of lettuce, no PCs were detected in the root given either Cd treatment.The high amounts of PCs produced in barley root could have contributed to preferential retention of Cd in barley roots.

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Section: Extraction Of Thiol-containing Moleculesmentioning

confidence: 99%

“…The thiol-containing compounds were derivatizated with MBrB following the procedures of Rijstenbil and Wijnholds (1996) and Sneller et al (2000), as described in Minocha et al (2008).…”

Section: Derivatization Of Thiol Groupsmentioning

confidence: 99%

Reduced translocation of cadmium from roots is associated with increased production of phytochelatins and their precursors

Akhter

McGarvey

Macfie

2012

Journal of Plant Physiology

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“…Thi ols were resolved and eluted at a flow rate of 1 mL min ¡1 and detected by fluo res cence (Jas co 821-FP Intel li gent Spec tro flu om e ter) with exci ta tion at 380 nm and emission at 480 nm (Klap heck, 1988;Snel ler et al, 2000). PC peaks were iden ti fied with syn the sized PC stan dards, as described in Lima et al (2006).…”

Section: Sep a Ra Tion And Deter MI Na Tion Of Thiol Com Poundsmentioning

confidence: 99%

Mercury intracellular partitioning and chelation in a salt marsh plant, Halimione portulacoides (L.) Aellen: Strategies underlying tolerance in environmental exposure

et al. 2009

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In the pres ence of metal stress, plants can resort to a series of tol er ance mech a nisms. There fore field studies should be under taken in order to eval u ate the real role of these mech a nisms in stress cop ing. The aim of this paper was to clar ify the bio chem i cal pro cesses behind mer cury tol er ance in Ha lim i one por tu la co ides (L.) Ael len (Caryo phyll ales: Cheno po di a ceae) col lected in a mer cury con tam i nated salt marsh. Dif fer ent frac tions of mer cury were sep a rated: buffer-sol u ble (mainly cyto solic) and insol u ble mer cury (mainly asso ci ated with mem branes and cell walls). The amounts in each frac tion of metal were com pared and related to metal dis tribu tion within plant organs. Pro tein-mer cury com plexes were iso lated and ana lysed for their thiol con tent in order to assess wether the tol er ance of this salt marsh plant was asso ci ated with the induc tion of metal che lation by phy to chel a tins. Over all, the mer cury tol er ance strat e gies of the plant are likely to involve root cell wall immo bi li za tion as a major mech a nism of metal resis tance, rather than metal che la tion in the cyto solic fraction. Nev er the less, phy to chel a tins were dem on strated to che late mer cury under envi ron men tal expo sure.

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“…Four-week-old plants were exposed to 20 M Cd 2ϩ in hydroponic medium for 72 h, and then tissue extracts were prepared by using glass homogenizers and thiols labeled by monobromobimane as described (9,24). Synthesized standards were used for the identification of PCs (␥-EC) 2 G (PC 2 ), (␥-EC) 3 G (PC 3 ), and (␥-EC) 4 G (PC 4 ).…”

Section: Plant Material Growth Conditions and Metal Stress Treatmentsmentioning

confidence: 99%

Long-distance root-to-shoot transport of phytochelatins and cadmium in Arabidopsis

Gong

Lee²,

Schroeder³

2003

Proc. Natl. Acad. Sci. U.S.A.

308

234

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Phytochelatin synthases (PCS) mediate cellular heavy-metal resistance in plants, fungi, and worms. However, phytochelatins (PCs) are generally considered to function as intracellular heavymetal detoxification mechanisms, and whether long-distance transport of PCs occurs during heavy-metal detoxification remains unknown. Here, wheat TaPCS1 cDNA expression was either targeted to Arabidopsis roots with the Arabidopsis alcohol dehydrogenase (Adh) promoter (Adh:: TaPCS1͞cad1 C admium is a widespread nonessential toxic heavy metal, released into the biosphere mostly by modern industry (1). Phytochelatins (PCs) play an essential role in heavy-metal detoxification in plants and fungi (2, 3). PCs chelate heavy metals and then PC-metal complexes are translocated across the tonoplast and sequestered in vacuoles (4, 5), thus decreasing the heavy-metal content in the cytosol of yeast and plant cells. PCs are synthesized from glutathione by the enzyme PC synthase (PCS) (6-9). However, PC research has focused mainly on PC transport into vacuoles (4, 5), and it remains unknown whether long-distance transport of PCs or PCS occurs in plants.Cd 2ϩ is taken up by plants, and plants have been proposed to provide an efficient system for heavy-metal removal from soils (10, 11). Soil composition affects Cd 2ϩ sensitivity. For example, silicon in soil reduces heavy-metal sensitivity (12, 13). Normally, Cd 2ϩ concentrations in roots are at least 10 times greater than those in shoots (14). However, for efficient phytoextraction from soils, heavy metals must be translocated into aerial tissues for later harvest. Translocation of Cd 2ϩ from roots to shoots has been studied in diverse plant species (15-18) and has been proposed to occur via the xylem of Indian mustard in a PCindependent manner (16). The molecular mechanisms for rootto-shoot Cd 2ϩ transport remain largely unknown, and PCs are predicted not to undergo long-distance transport but to enhance vacuolar heavy-metal sequestration in the cells in which they are produced.The cad1-3 mutant is a recessive, loss-of-function mutation in the Arabidopsis AtPCS1 gene (7) (9) by PCR. The TaPCS1 PCR fragment was subcloned into pBluescript II SK(ϩ) vector. A 3ϫ myc tag DNA sequence was constructed by PCR recovery from a plasmid containing c-myc and then subcloned into the pGEM-T Easy vector (Promega). The subcloned myc fragment was recovered with SpeI and SacI enzymes and then fused to the 3Ј end of the TaPCS1 ORF in the pBluescript II SK(ϩ) vector at the SpeI site. All PCR products were confirmed by sequencing (Retrogen, La Jolla, CA). The fusion sequence was then digested with BamHI and SacI and subcloned into the binary expression vector pBI121, either with the cauliflower mosaic virus 35S or the Arabidopsis alcohol dehydrogenase (Adh) promoter (20). Both constructs were transformed into the PC-deficient Arabidopsis mutant cad1-3 by direct Agrobacterium tumefaciens-mediated transformation using the floral dip technique (21). T2 seeds with 3:1 segregation on kanamycin plates were used for hom...

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Derivatization of Phytochelatins from Silene vulgaris, Induced upon Exposure to Arsenate and Cadmium: Comparison of Derivatization with Ellman's Reagent and Monobromobimane

Cited by 173 publications

References 27 publications

Reduced translocation of cadmium from roots is associated with increased production of phytochelatins and their precursors

Reduced translocation of cadmium from roots is associated with increased production of phytochelatins and their precursors

Mercury intracellular partitioning and chelation in a salt marsh plant, Halimione portulacoides (L.) Aellen: Strategies underlying tolerance in environmental exposure

Long-distance root-to-shoot transport of phytochelatins and cadmium in Arabidopsis

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