A novel plant glutathione S-transferase/peroxidase suppresses Bax lethality in yeast

The mammalian inducer of apoptosis Bar is lethal when expressed in yeast an d plant cells. To identify potential inhibitors of Bax in plants we transfo rmed yeast cells expressing Bax with a tomato cDNA library and we selected for cells surviving after the induction of Bax. This genetic screen allows for the identification of plant genes, which inhibit either directly or ind irectly the lethal phenotype of Bax. Using this method a number of cDNA clo nes were isolated, the more potent of which encodes a protein homologous to the class theta glutathione S-transferases. This Bax-inhibiting (BI) prote in was expressed in Escherichia coli and found to possess glutathione S-tra nsferase (GST) and weak glutathione peroxidase (GPX) activity. Expression of Bax in yeast decreases the intracellular levels of total glut athione, causes a substantial reduction of total cellular phospholipids, di minishes the mitochondrial membrane potential, and alters the intracellular redox potential. Co-expression of the BI-GST/GPX protein brought the total glutathione levels back to normal and re-established the mitochondrial mem brane potential but had no effect on the phospholipid alterations. Moreover , expression of BI-GST/GPX in yeast was found to significantly enhance resi stance to H2O2-induced stress. These results underline the relationship bet ween oxidative stress and Bax-induced death in yeast cells and demonstrate that the yeast-based genetic strategy described here is a powerful tool for the isolation of novel antioxidant and anti-apoptotic genes.