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J. Biol. Chem. 275 (38): 29207-29216

© 2000 by The American Society for Biochemistry and Molecular Biology, Inc.

A Novel Plant Glutathione S-Transferase/Peroxidase Suppresses Bax Lethality in Yeast*

Sotirios C. Kampranis{ddagger}§, Radostina Damianova{ddagger}§, Mirna Atallah{ddagger}§, Garabet Toby, Greta Kondi{ddagger}, Philip N. Tsichlis||, , and Antonios M. Makris{ddagger}**

From the {ddagger}Mediterranean Agronomic Institute of Chania, Chania 73100, Greece, Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, and ||Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, Pennsylvania 19107

ABSTRACT Back to Top

Abstract: The mammalian inducer of apoptosis Bax is lethal when expressed in yeast and plant cells. To identify potential inhibitors of Bax in plants we transformed 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 indirectly the lethal phenotype of Bax. Using this method a number of cDNA clones were isolated, the more potent of which encodes a protein homologous to the class {theta} glutathioneS-transferases. This Bax-inhibiting (BI) protein was expressed in Escherichia coli and found to possess glutathione S-transferase (GST) and weak glutathione peroxidase (GPX) activity. Expression of Bax in yeast decreases the intracellular levels of total glutathione, causes a substantial reduction of total cellular phospholipids, diminishes 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 membrane potential but had no effect on the phospholipid alterations. Moreover, expression of BI-GST/GPX in yeast was found to significantly enhance resistance to H2O2-induced stress. These results underline the relationship between 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 antiapoptotic genes.

Received for publication March 21, 2000. Revision received May 22, 2000.

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