A novel plant glutathione S-transferase/peroxidase suppresses Bax lethality in yeast
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A novel plant glutathione S-transferase/peroxidase suppresses Bax lethality in yeast. / Kampranis, S C; Damianova, R; Atallah, M; Toby, G; Kondi, G; Tsichlis, P N; Makris, A M.
In: The Journal of Biological Chemistry, Vol. 275, No. 38, 22.09.2000, p. 29207-16.Research output: Contribution to journal › Journal article › Research › peer-review
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TY - JOUR
T1 - A novel plant glutathione S-transferase/peroxidase suppresses Bax lethality in yeast
AU - Kampranis, S C
AU - Damianova, R
AU - Atallah, M
AU - Toby, G
AU - Kondi, G
AU - Tsichlis, P N
AU - Makris, A M
PY - 2000/9/22
Y1 - 2000/9/22
N2 - 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 glutathione S-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 H(2)O(2)-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.
AB - 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 glutathione S-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 H(2)O(2)-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.
KW - Amino Acid Sequence
KW - Apoptosis
KW - Cell Survival
KW - Gene Expression Regulation, Fungal
KW - Glutathione Transferase
KW - Molecular Sequence Data
KW - Oxidative Stress
KW - Plant Proteins
KW - Proto-Oncogene Proteins
KW - Proto-Oncogene Proteins c-bcl-2
KW - Saccharomyces cerevisiae
KW - Sequence Alignment
KW - bcl-2-Associated X Protein
U2 - 10.1074/jbc.M002359200
DO - 10.1074/jbc.M002359200
M3 - Journal article
C2 - 10859306
VL - 275
SP - 29207
EP - 29216
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 38
ER -
ID: 159085426