THE INTRACELLULAR LOCATION OF NADH-CYTOCHROME B(5) REDUCTASE MODULATES THE CYTOTOXICITY OF THE MITOMYCINS TO CHINESE-HAMSTER OVARY CELLS

NADH:cytochrome b(5) reductase activates the mitomycins to alkylating intermediates in vitro. To investigate the intracellular role of this enzyme in mitomycin bioactivation, Chinese hamster ovary cell transfec tants overexpressing rat NADH:cytochrome b(5) reductase were generated . An NADH:cytochrome b(5) reductase-transfected clone expressed 9-fold more enzyme than did parental cells; the levels of other mitomycin-ac tivating oxidoreductases were unchanged. Although this enzyme activate s the mitomycins in vitro, its overexpression in living cells caused d ecreases in sensitivity to mitomycin C in air and decreases in sensiti vity to porfiromycin under both air and hypoxia, Mitomycin C cytotoxic ity under hypoxia was similar to parental cells. Because NADH:cytochro me b(5) reductase resides predominantly in the mitochondria of these c ells, this enzyme may sequester these drugs in this compartment, there by decreasing nuclear DNA alkylations and reducing cytotoxicity. A cyt osolic form of NADH:cytochrome b(5) reductase was generated. Transfect ants expressing the cytosolic enzyme were restored to parental line se nsitivity to both mitomycin C and porfiromycin in air with marked incr eases in drug sensitivity under hypoxia. The results implicate NADH:cy tochrome b(5) reductase in the differential bioactivation of the mitom ycins and indicate that the subcellular site of drug activation can ha ve complex effects on drug cytotoxicity.