Ectopic expression γ-glutamylcysteine synthetase of Vicia sativa increased cadmium tolerance in Arabidopsis.

Glutathione (GSH) is a multifunctional essential biothiol, and its metabolism is important for plant against toxic metals and metalloids. γ-Glutamylcysteine (γ-EC), which is catalyzed by γ-Glutamylcysteine synthetase (γ-ECS), is a rate-limiting intermediate in GSH synthesis. Here, a γ-ECS gene (Vsγ-ECS) from Vicia sativa was cloned, and its function in modulating Cd tolerance was studied. Vsγ-ECS is a chloroplast localization protein, and the expression of Vsγ-ECS was upregulated by Cd stress in root of V. sativa. Heterologous expression of Vsγ-ECS (35S::Vsγ-ECS) in Arabidopsis enhanced the Cd tolerance of plants through improved primary root length, fresh weight, chlorophyll content and low degree of oxidation associated with reduced H2O2 and lipid peroxidation. However, the Cd accumulation of Arabidopsis had no effect on Vsγ-ECS overexpression. Further analysis showed that the increased Cd tolerance in 35S::Vsγ-ECS was mainly due to the capacity of increasing GSH synthesis that improved Cd chelation by GSH and phytochelatins (PCs) and alleviated the oxidative stress caused by Cd stress. In summary, a γ-ECS was characterized from V. sativa, and it demonstrated a property for increasing GSH and PC synthesis to protect plants from Cd poisoning.

Specific accumulation of CYP94A1 transcripts after exposure to gaseous benzaldehyde: induction of lauric acid ω-hydroxylase activity in Vicia sativa exposed to atmospheric pollutants.

The effects of air pollutants such as aldehydes, ozone, nitrogen dioxide and benzene on fatty acid ω-hydroxylase activity in Vicia sativa microsomes have been investigated. Four days old etiolated V. sativa seedlings were exposed to different concentrations of selected pollutants for varying exposure times. Growing etiolated V. sativa seedlings in air containing the gaseous benzaldehyde (150 nM) led to an 8-fold enhancement of lauric acid ω-hydroxylase activity in microsomes of treated plants compared to controls grown in pure air (96 ± 10 versus 12 ± 2 pmol/min/mg protein, respectively). The induction increased with increasing gas phase concentrations (10-1300 nM) and the maximum of activity was measured after 48 h of exposure. Northern blot analysis revealed that this induction occurred via transcriptional activation of the gene coding for CYP94A1. The absence of CYP94A2 and CYP94A3 transcription activation together with the missing effect on epoxide hydrolases activities indicate the specificity of CYP94A1 induction by benzaldehyde. Exposure to nitrogen dioxide, ozone and formaldehyde also stimulated lauric acid ω-hydroxylases activity while exposure to benzene did not show any effect.

Enzymatic synthesis of (R)-cyanohydrins by a novel (R)-oxynitrilase from Vicia sativa L.

The defatted seed meal of common vetch (Vicia sativa L.) served as a source of (R)-oxynitrilase which catalyzed the enantioselective addition of HCN to aromatic, heteroaromatic, fluoro-substituted aromatic aldehydes to produce the corresponding enantiomeric pure cyanohydrins in yields of 52-100% and 88-99% ee at 12 degrees C under micro-aqueous medium (diisopropyl ether) without addition of buffer solution.

CYP94A1, a plant cytochrome P450-catalyzing fatty acid omega-hydroxylase, is selectively induced by chemical stress in Vicia sativa seedlings.

CYP94A1 is a cytochrome P450 (P450) catalyzing fatty acid (FA) omega-hydroxylation in Vicia sativa seedlings. To study the physiological role of this FA monooxygenase, we report here on its regulation at the transcriptional level (Northern blot). Transcripts of CYP94A1, as those of two other P450-dependent FA hydroxylases (CYP94A2 and CYP94A3) from V. sativa, are barely detectable during the early development of the seedlings. CYP94A1 transcripts, in contrast to those of the two other isoforms, are rapidly (less than 20 min) and strongly (more than 100 times) enhanced after treatment by clofibrate, an hypolipidemic drug in animals and an antiauxin (p-chlorophenoxyisobutyric acid) in plants, by auxins (2,4-dichlorophenoxyacetic acid and indole-3-acetic acid), by an inactive auxin analog (2,3-dichlorophenoxyacetic acid), and also by salicylic acid. All these compounds activate CYP94A1 transcription only at high concentrations (50-500 microM range). In parallel, these high levels of clofibrate and auxins modify seedling growth and development. Therefore, the expression of CYP94A1 under these conditions and the concomitant morphological and cytological modifications would suggest the implication of this P450 in a process of plant defense against chemical injury.

The activity and location of cathepsin D inhibitor in seeds of common vetch (Vicia sativa L.).

The activity of cathepsin D inhibitor is markedly higher in common vetch seed coat than in embryo cotyledons. The occurrence of considerable amounts of the inhibitor in the seed coat of vetch was confirmed by the fluorescent microscopic technique, with the use of fluorescein-marked cathepsin D.