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. 2004 May;135(1):377-83.
doi: 10.1104/pp.103.026989. Epub 2003 Dec 11.

Overexpression of selenocysteine methyltransferase in Arabidopsis and Indian mustard increases selenium tolerance and accumulation

Danika L LeDuc  1 , Alice S Tarun, Maria Montes-Bayon, Juris Meija, Michele F Malit, Carol P Wu, Manal AbdelSamie, Chih-Yuan Chiang, Abderrhamane Tagmount, Mark deSouza, Bernhard Neuhierl, August Böck, Joseph Caruso, Norman Terry
Affiliations

Overexpression of selenocysteine methyltransferase in Arabidopsis and Indian mustard increases selenium tolerance and accumulation

Danika L LeDuc et al. Plant Physiol. 2004 May.

Abstract

A major goal of phytoremediation is to transform fast-growing plants with genes from plant species that hyperaccumulate toxic trace elements. We overexpressed the gene encoding selenocysteine methyltransferase (SMT) from the selenium (Se) hyperaccumulator Astragalus bisulcatus in Arabidopsis and Indian mustard (Brassica juncea). SMT detoxifies selenocysteine by methylating it to methylselenocysteine, a nonprotein amino acid, thereby diminishing the toxic misincorporation of Se into protein. Our Indian mustard transgenic plants accumulated more Se in the form of methylselenocysteine than the wild type. SMT transgenic seedlings tolerated Se, particularly selenite, significantly better than the wild type, producing 3- to 7-fold greater biomass and 3-fold longer root lengths. Moreover, SMT plants had significantly increased Se accumulation and volatilization. This is the first study, to our knowledge, in which a fast-growing plant was genetically engineered to overexpress a gene from a hyperaccumulator in order to increase phytoremediation potential.

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Figures

Figure 1.
Figure 1.
A schematic representation of key metabolic intermediates of Se in nonaccumulators and hyperaccumulators. Reactions that may not be specific to hyperaccumulators convert MetSeCys, the end product of the hyperaccumulator pathway, to DMDSe, a volatile gas.
Figure 2.
Figure 2.
Molecular analysis of SMT transgenic plants. A, Northern-blot analysis of equimolar total RNA isolated from wild-type and SMT Arabidopsis probed with the SMT gene. B, RT-PCR analysis of cDNA derived from equimolar amounts of total RNA isolated from wild-type and SMT Indian mustard using PCR primers directed toward the SMT gene (shown in inverse). Immunoblot of equimolar amounts of protein isolated from wild-type and SMT Arabidopsis (C) and SMT Indian mustard (D) using an antibody specific for the A. bisulcatus SMT gene. The strong top band in the Arabidopsis immunoblot and the faint top band in the Indian mustard immunoblot correspond to a native protein, which has cross-reactivity with the SMT antibody. The SMT antibody did not bind to any protein At7-23. This line was excluded from further study.
Figure 3.
Figure 3.
Seedling tolerance experiments with SMT transgenic plants. A, Fresh weight and root length Tolerance Index for wild-type and SMT (lines 8–14 and 34–10) Arabidopsis seedlings treated with 25 μm selenite or SeCys. The Tolerance Index is calculated by dividing treatment values with the average corresponding control values for that line. B, Fresh weights and root lengths of wild-type and SMT (lines 3–11 and 5–13) Indian mustard seedlings supplied with no selenium or 200 μm selenate or selenite. Experiments with other selenocompounds were conducted, but only statistically significant data are shown. Values for SMT transgenic lines that are statistically significantly different from wild-type values are denoted with either an a (P < 0.01) or b (P < 0.05).
Figure 4.
Figure 4.
Se accumulation in Arabidopsis seedlings treated with 25 μm selenate (A); Indian mustard seedlings treated with 200 μm selenate (B); Indian mustard seedlings treated with 100 μm selenite (C); and mature Indian mustard treated with 50 μm selenite (D). Experiments with other selenocompounds were conducted, but only statistically significant data are shown. Values for SMT transgenic lines that are statistically significantly different from wild-type values are denoted with either an a (P < 0.01) or b (P < 0.05).
Figure 5.
Figure 5.
MetSeCys concentrations in leaves of mature Indian mustard plants treated with 50 μm selenate or 50 μm selenite (A). Values for SMT transgenic lines that are statistically significantly different from wild-type values are denoted with an a (P < 0.01) or b (P < 0.05). The values are calculated on a per fresh weight basis. B, The average percentage of HCl-extractable Se in the form of MetSeCys in mature Indian mustard leaves treated with selenate or selenite.
Figure 6.
Figure 6.
Se volatilization in Arabidopsis mature plants treated with 25 μm SeCys (A) and mature Indian mustard treated with 20 μm selenate (B). Values for SMT transgenic lines that are statistically significantly different from wild-type values are denoted with either an a (P < 0.01) or b (P < 0.05).
Figure 7.
Figure 7.
Amount of DMDSe present in the headspace of wild-type and SMT Indian mustard seedlings contained in airtight serum bottles and treated with 200 μm concentrations of selenate or selenite for 8 d. The gases were analyzed by GC-ICP-MS. The values are those calculated on a per gram dry weight basis.
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