Search

A group of selenium (Se)-hyperaccumulating species belonging to the genus Astragalus are known for their capacity to accumulate up to 0.6% of their foliar dry weight as Se, with most of this Se being in the form of Se-methylselenocysteine (MeSeCys). Here, we report the isolation and molecular characterization of the gene that encodes a putative selenocysteine methyltransferase (SMT) enzyme from the non-accumulator Astragalus drummondii and biochemically compare it with an authentic SMT enzyme from the Se-hyperaccumulator Astragalus bisulcatus, a related species that lives within the same native habitat. The non-accumulator enzyme (AdSMT) shows a high degree of homology with the accumulator enzyme (AbSMT) but lacks the selenocysteine methyltransferase activity in vitro, explaining why little or no detectable levels of MeSeCys accumulation are observed in the non-accumulator plant. The insertion of mutations on the coding region of the non-accumulator AdSMT enzyme to better resemble enzymes that originate from Se accumulator species results in increased selenocysteine methyltransferase activity, but these mutations were not sufficient to fully gain the activity observed in the AbSMT accumulator enzyme. We demonstrate that SMT is localized predominantly within the chloroplast in Astragalus, the principal site of Se assimilation in plants. By using a site-directed mutagenesis approach, we show that an Ala to Thr amino acid mutation at the predicted active site of AbSMT results in a new enzymatic capacity to methylate homocysteine. The mutated AbSMT enzyme exhibited a sixfold higher capacity to methylate selenocysteine, thereby establishing the evolutionary relationship of SMT and homocysteine methyltransferase enzymes in plants.

Several Astragalus species have the ability to hyperaccumulate selenium (Se) when growing in their native habitat. Given that the biochemical properties of Se parallel those of sulfur (S), we examined the activity of key S assimilatory enzymes ATP sulfurylase (ATPS), APS reductase (APR), and serine acetyltransferase (SAT), as well as selenocysteine methyltransferase (SMT), in eight Astragalus species with varying abilities to accumulate Se. Se hyperaccumulation was found to positively correlate with shoot accumulation of S-methylcysteine (MeCys) and Se-methylselenocysteine (MeSeCys), in addition to the level of SMT enzymatic activity. However, no correlation was observed between Se hyperaccumulation and ATPS, APR, and SAT activities in shoot tissue. Transgenic Arabidopsis thaliana overexpressing both ATPS and APR had a significant enhancement of selenate reduction as a proportion of total Se, whereas SAT overexpression resulted in only a slight increase in selenate reduction to organic forms. In general, total Se accumulation in shoots was lower in the transgenic plants overexpressing ATPS, PaAPR, and SAT. Root growth was adversely affected by selenate treatment in both ATPS and SAT overexpressors and less so in the PaAPR transgenic plants. Such observations support our conclusions that ATPS and APR are major contributors of selenate reduction in planta. However, Se hyperaccumulation in Astragalus is not driven by an overall increase in the capacity of these enzymes, but rather by either an increased Se flux through the S assimilatory pathway, generated by the biosynthesis of the sink metabolites MeCys or MeSeCys, or through an as yet unidentified Se assimilation pathway.

This study presents the effects of methyl jasmonate (MeJA) on growth, N uptake, N partitioning, and N storage in taproots of non-nodulated alfalfa (cv. Lodi). When compared to untreated plants, addition of 100 µM MeJA to the nutrient solution for 14 days reduced total growth and modified biomass partitioning between shoots and roots in favour of taproots and lateral roots. MeJA decreased N uptake (after 7 days) and increased N partitioning towards roots after 14 days. This preferential N partitioning to roots was accompanied by increased N storage in taproots as soluble proteins. Compared to total soluble proteins, VSP accumulation occurred earlier (7 days), and was greater (2-fold increase) in plants treated with 100 µ MMeJA. Steady-state transcript levels for two VSPs (32 and 57 kDa) also increased markedly (about 4-fold) in roots of plants treated with 100 µM MeJA. This suggests that MeJA could act directly (transcriptional regulation) or indirectly (via the changes of N partitioning among alfalfa organs) on N storage as soluble proteins and in particular, VSPs. Because the deduced amino acid sequence of the 32 kDa VSP clone reveals high homology with Class III chitinases, we propose that the 32 kDa VSP may have a role in pathogen defense, in addition to its function as a storage protein. [ABSTRACT FROM AUTHOR]