Abstract
Regulation of the expression of the cytosolic O-acetylserine(thiol)lyase gene (Atcys-3A) from Arabidopsis thaliana under heavy metal stress conditions has been investigated. Northern blot analysis of Atcys-3A expression shows a 7-fold induction after 18 h of cadmium treatment. Addition of 50 microm CdCl(2) to the irrigation medium of mature Arabidopsis plants induces a rapid accumulation of the mRNA throughout the leaf lamina, the root and stem cortex, and stem vascular tissues when compared with untreated plants, as observed by in situ hybridization. High pressure liquid chromatography analysis of GSH content shows a transient increase after 18 h of metal treatment. Our results are compatible with a high cysteine biosynthesis rate under heavy metal stress required for the synthesis of GSH and phytochelatins, which are involved in the plant detoxification mechanism. Arabidopsis-transformed plants overexpressing the Atcys-3A gene by up to 9-fold show increased tolerance to cadmium when grown in medium containing 250 microm CdCl(2), suggesting that increased cysteine availability is responsible for cadmium tolerance. In agreement with these results, exogenous addition of cystine can, to some extent, also favor the growth of wild-type plants in cadmium-containing medium. Cadmium accumulates to higher levels in leaves of tolerant transformed lines than in wild-type plants.
Highlights
Back control of the ␥-glutamylcysteine synthetase by GSH
Our results are compatible with a high cysteine biosynthesis rate under heavy metal stress required for the synthesis of GSH and phytochelatins, which are involved in the plant detoxification mechanism
The last step of cysteine biosynthesis is catalyzed by the O-acetylserine(thiol)lyase enzyme (OASTL)1 that incorporates sulfide into the O-acetyl-L-serine molecule
Summary
Plant Material, Growth, and Treatments—Wild-type A. thaliana (ecotype Columbia) plants were grown on moist vermiculite supplemented with Hoagland medium at 20 °C in the light and 18 °C in the dark, under a 16-h white light/8-h dark photoperiod. Cadmium chloride treatments were performed by addition to the Hoagland medium of CdCl2 to 50 M final concentration, unless otherwise indicated. Seeds were surface-sterilized and germinated on solid MS medium with and without CdCl2 at indicated concentrations, after cold treatment for 1 day to improve germination. For circadian experiments plants were grown on soil under a 16-h white light/8-h dark photoperiod for 3 weeks, before harvesting every 4 h for 36 h. After this treatment, plants were shifted to constant light and harvested every 4 h during 1 day.
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