Glutathione S -transferases in Arabidopsis and their regulation by herbicide safeners

Benjamin P DeRidder, Purdue University

Abstract

Glutathione S-transferases (GSTs) in plants have been implicated in the detoxification of agrochemicals such as herbicides. Herbicide safeners are compounds that enhance the herbicide-detoxifying activity of GSTs and other proteins in grass crops but not in target weeds. Little is known about how safeners induce detoxification systems in plants. In this study, Arabidopsis was evaluated as a model to study safener response mechanisms in higher plants, in an attempt to understand the basis for GST regulation by safeners. In the current study, several safeners were shown to induce the expression of specific Arabidopsis GSTs at the RNA and protein levels, and to enhance GST activity against herbicides and model substrates. However, all safeners did not induce the same profile of GSTs in Arabidopsis, suggesting multiple pathways are involved in the regulation of GST expression by safeners. When GSTs from safener-treated plants were purified by affinity chromatography and displayed by 2-D SDS-PAGE, a 26 kD protein that was induced by a number of safeners was identified as a novel GST, AtGSTU19. When expressed in E. coli, AtGSTU19 had high activity with model substrates and conjugated several chloroacetamide herbicides to GSH at rates similar to those reported for herbicide-metabolizing GSTs in sorghum and maize. Based on its activity and abundance in protein extracts, it is likely that AtGSTU19 accounts for a large proportion of the increase in GST activity following safener treatment. Levels of AtGSTU19 RNA and protein increased predominantly in roots in response to safeners that are used in cereal crops to reduce chloroacetamide injury. The tissue-specific expression of AtGSTU19 was examined in vivo using transgenic lines that carrying a GFP reporter gene driven by the AtGSTU19 promoter. Low basal expression of the reporter in roots was observed in untreated plants, while expression was induced moderately in shoots and highly in roots of safener-treated plants. The effect of overexpressing AtGSTU19 in Arabidopsis on herbicide tolerance was investigated. The results suggest that localization of GST expression to certain plant tissues is an important factor determining safener efficacy. Finally, a proteomic analysis of the Arabidopsis GST family examined the expression of eight abundant GSTs, including AtGSTU19, before and after safener treatment. MS-MS was used to identify differentially-labeled tryptic peptides derived from GST proteins in control and safener-treated plants. Differences in the abundance of these peptides indicated that expression of specific GSTs changed in response to safener-treatment. This work establishes Arabidopsis as a model to study the regulation of GSTs by safeners, and offers an explanation for differences in safener action between cereal crops and weeds.

Degree

Ph.D.

Advisors

Goldsbrough, Purdue University.

Subject Area

Molecular biology|Botany|Genetics

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