The hypersensitivity of the Chinese hamster ovary variant BL-10 to bleomycin killing is due to a lack of glutathione S-transferase-alpha activity.
As a means to understand the fundamental mechanisms of bleomycin cell killing, we previously isolated 19 bleomycin-sensitive mutants which represent at least six genetically distinct complementation groups (T.D. Stamato, B. Peters, P. Patil, N. Denko, R. Weinstein, and A. Giaccia. Cancer Res., 47: 1588-1592, 1987). One class of mutants represented by the cell line BL-10 displays only hypersensitivity to killing by bleomycin in both acute (16 h) and chronic treatments but no sensitivity to killing by other DNA-damaging agents. Complementation studies between this mutant and human fibroblasts suggested that the human gene which corrects the defect of BL-10 rested on human chromosome 6. It has been reported that the gene for human glutathione S-transferase (GST) alpha also resides on chromosome 6. Measurements of selenium-independent peroxidase (alpha-GST + glutathione peroxidase) activity in wild-type Chinese hamster ovary (CHO) cells, using cumene hydrogen peroxide as a substrate, gave a value of 112 nmol of glutathione oxidized/min/mg protein compared with 88.1 nmol of glutathione oxidized/min/mg protein for BL-10. Measurement of the selenium-dependent peroxidase activity, using H2O2 as a substrate, resulted in 65.9 nmol of reduced glutathione oxidized/min/mg protein in CHO and 81.5 nmol of reduced glutathione oxidized/min/mg protein for BL-10. In other words, BL-10 cells did not exhibit a difference in their ability to metabolize both substrates in contrast to CHO cells. This indicates that BL-10 possesses little alpha-GST activity. Transfection of BL-10 cells with a mammalian expression vector containing the alpha-GST gene increases the survival of BL-10 to bleomycin and does not increase the bleomycin resistance of two other bleomycin mutants which lie in different genetic complementation groups. These data strongly implicate a role for alpha-GST in the resistance of cells to bleomycin.
Giaccia
A J
AJ
Department of Radiation Oncology, Stanford University School of Medicine, California 94305.
Lewis
A D
AD
Denko
N C
NC
Cholon
A
A
Evans
J W
JW
Waldren
C A
CA
Stamato
T D
TD
Brown
J M
JM
D000818:Animals; D001761:Bleomycin / Q000494:pharmacology*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects*; D002871:Chromosome Banding; D002896:Chromosomes, Human, Pair 6; D002999:Clone Cells; D006224:Cricetinae; D003412:Cricetulus; D004305:Dose-Response Relationship, Drug; D004351:Drug Resistance / Q000235:genetics; D005260:Female; D005816:Genetic Complementation Test; D014644:Genetic Variation; D005982:Glutathione Transferase / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D006822:Hybrid Cells / Q000166:cytology* / Q000187:drug effects; D010053:Ovary
D001761:Bleomycin; D005982:Glutathione Transferase