Exploiting plants for glutathione (GSH) production: Uncoupling GSH synthesis from cellular controls results in unprecedented GSH accumulation.

Glutathione (GSH) is a key factor for cellular redox homeostasis and tolerance against abiotic and biotic stress (May et al., 1998; Noctor et al., 1998a). Previous attempts to increase GSH content in plants have met with moderate success (Rennenberg et al., 2007), largely because of tight and multilevel control of its biosynthesis (Rausch et al., 2007). Here, we report the in planta expression of the bifunctional gamma-glutamylcysteine ligase-glutathione synthetase enzyme from Streptococcus thermophilus (StGCL-GS), which is shown to be neither redox-regulated nor sensitive to feedback inhibition by GSH. Transgenic tobacco plants expressing StGCL-GS under control of a constitutive promoter reveal an extreme accumulation of GSH in their leaves (up to 12 micromol GSH/gFW, depending on the developmental stage), which is more than 20- to 30-fold above the levels observed in wild-type (wt) plants and which can be even further increased by additional sulphate fertilization. Surprisingly, this dramatically increased GSH production has no impact on plant growth while enhancing plant tolerance to abiotic stress. Furthermore, StGCL-GS-expressing plants are a novel, cost-saving source for GSH production, being competitive with current yeast-based systems (Li et al., 2004).

Expression of BjMT2, a metallothionein 2 from Brassica juncea, increases copper and cadmium tolerance in Escherichia coli and Arabidopsis thaliana, but inhibits root elongation in Arabidopsis thaliana seedlings.

The protective function of a plant type-2 metallothionein was analysed after expression in Escherichia coli and in Arabidopsis thaliana seedlings. BjMT2 from Brassica juncea was expressed in E. coli as a TrxABjMT2 fusion protein. After affinity chromatography and cleavage from the TrxA domain, pure BjMT2 protein was obtained which strongly reacted with the thiol reagent monobromobimane. Escherichia coli cells expressing the TrxABjMT2 fusion were more tolerant to Cu2+ and Cd2+ exposure than control strains. Likewise, when BjMT2 cDNA was expressed in A. thaliana under the regulation of the 35S promoter, seedlings exhibited an increased tolerance against Cu2+ and Cd2+ based on shoot growth and chlorophyll content. Analysis of transiently transformed cells of A. thaliana and tobacco leaves by confocal laser scanning microscopy (CLSM) revealed exclusive cytosolic localization of a BjMT2::EGFP (enhanced green fluorescent protein) fusion protein in control and heavy metal-exposed plant cells. Remarkably, ectopic expression of BjMT2 reduced root growth in the absence of heavy metal exposure, whereas in the presence of 50 or 100 microM Cu2+ root growth in control and transgenic lines was identical. The results indicate that in A. thaliana, root and shoot development are differentially affected by ectopic expression of BjMT2.