Regulation of an Arabidopsis glutathione S-transferase gene (AtGSTU19) by herbicide safeners

Nahla A El-Sherif, Purdue University

Abstract

Herbicide safeners are interesting chemical compounds that enhance the selectivity of herbicides in cereal crops. The most widely accepted explanation for how safeners increase herbicide tolerance is that they induce a variety of herbicide detoxification activities in the crop plant. Among the enzymes that are induced by safeners are glutathione S-transferases (GSTs), which are critical components in phase II of the detoxification system. Plant GSTs have well described roles in the metabolism of many herbicides in crops. However, individual GSTs are also expressed in response to many other cues, including other forms of biotic and abiotic stress, suggesting that GSTs also have diverse functions in normal metabolism. One question about safeners that remains unanswered is how they induce multiple components of the xenobiotic detoxification system in plants. Previous studies have shown that AtGSTU19 is among the most highly induced Arabidopsis GST genes. Safeners also increased the concentration of GSH and the expression of genes encoding glutathione-conjugate transporters. While safeners were able to increase the activity of multiple components of the detoxification system, they were unable to protect Arabidopsis plants from various chloroacetamide herbicides. Experiments with transgenic plants have shown that safeners fail to protect Arabidopsis from herbicide injury because the GSTs induced by these compounds are inefficient at conjugating chloroacetamide herbicides and are not expressed in the correct tissues. In spite of the fact that safeners fail to protect Arabidopsis, they do induce GST expression and this has allowed me to examine the mechanism by which herbicide safeners regulate the expression of AtGSTU19. A deletion analysis of the AtGSTU19 promoter showed that an as-1 element located approximately 150 bp upstream of the transcription start site is required for safener-induced expression of this gene. Specific mutations within the as-1 element also abolished the capacity of the AtGSTU19 promoter to respond to safeners. The as-1 promoter element therefore plays a critical role in AtSGTU19 expression and induction by safeners. Salicylic acid (SA) is known to regulate the expression of many genes that contain as-1 promoter elements. RNA expression of AtGSTU19 was induced by treatment of seedlings with SA, whereas benoxacor and fenclorim were unable to induce AtGSTU19 in SA-deficient NahG plants that express a bacterial SA hydroxylase gene. However, the sid2 mutant, which is unable to synthesize SA via the isochorismate synthase pathway, showed normal induction of AtGSTU19 by benoxacor. These results support the hypothesis that safeners function through a SA signaling pathway in Arabidopsis independent of SID2. To confirm that SA is required for safener activity and to investigate which SA biosynthesis pathway is involved in this process, we measured SA in Arabidopsis plants under control conditions and safener treatment. The safener benoxacor produced a substantial increase in SA concentration in Arabidopsis plants within 6 hours of treatment in both wild type and sid2 mutant. These results indicate the involvement of an SID2-independent SA biosynthesis pathway, from phenylalanine and benzoic acid, in activating the expression of AtGSTU19 in response to safeners. Further experiments will be required to determine if other components of the detoxification system are regulated in the same way by safeners. In conclusion, I have shown that SA is critical for the regulation of GST gene expression in response to safeners in Arabidopsis.

Degree

Ph.D.

Advisors

Goldsbrough, Purdue University.

Subject Area

Molecular biology|Plant sciences

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