Localization of glutathione-s-transferase and its effect on seedling development

Abstract

Plants have evolved various defense mechanisms that include antioxidants (easily oxidizable compounds of small molecular weight) and antioxidant enzymes to combat the effects of reactive oxygen species produced by abiotic and biotic stress. One such enzyme family is glutathione-S-transferase (GST). These enzymes detoxify various electrophilic and hydrophobic compounds via conjugation to the thiol group of glutathione (GSH), an antioxidant molecule that is abundantly present in plant cells. Tobacco plants transformed with NtI07, a cDNA clone that encodes a GST with glutathione peroxidase activity, have been shown to have increased tolerance to salt and temperature stress, as well as to have increased levels of oxidized glutathione (Roxas et al., 1997). Although GST has been established as an effective enzyme to protect seedlings under stressful conditions, exactly how the plant system is affected remains to be completely understood.

We have attempted to examine the effects of glutathione on seedling growth by using the specific glutathione synthesis inhibitor, buthionine sulfoximine (BSO). Results indicated that glutathione is required for normal seedling growth, and seedlings treated with BSO had higher lipid peroxidation levels than controls. Although we observed that exogenous glutathione also had an inhibitory effect on growth, seedlings treated with glutathione did not show higher levels of lipid peroxidation.

Seed storage protein mobilization patterns in GST-overexpressing and wildtype seedlings have also been investigated under normal and stress conditions. We found that GST-overexpressing seedlings mobilized their storage proteins more slowly than did wildtype under normal conditions, however the mobilization in the transgenic seedlings was less affected under salt stress than it was in wildtype.

Knowledge about the localized activity of GST could also help elucidate the mechanism of stress resistance. We used polyclonal antibodies raised against the protein to localize GST in two-week old tobacco seedlings and found the primary localization to be in the nucleus of both transgenic and wildtype seedlings, indicating that GST may be involved in protection against DNA damage.