Although an apparently generalized defect of
cytochrome c oxidase (COX) occurs in many patients with
subacute necrotizing encephalomyelopathy (Leigh's syndrome), the mode of inheritance in this disorder is not known. We transformed COX-deficient fibroblasts from a child with Leigh's syndrome with simian virus 40 to obtain cells with an infinite life span. These cells were still COX-deficient, grew normally in HAT medium, and were
ouabain-sensitive. We fused these cells with a HAT-sensitive,
ouabain-resistant variant of HeLa cells (HeLacot) and isolated surviving hybrid clones in
ouabain-containing HAT medium. Prolonged cultivation of the hybrids was accompanied by preferential loss of HeLacot
mitochondrial DNA (
mtDNA), as determined by
mtDNA restriction patterns of parental and hybrid cell
DNA with the
restriction endonuclease HaeII. COX activity was normal or higher than normal in hybrids, including the progeny of cell clones that had lost almost all the HeLacot
mtDNA. These data demonstrate that
COX deficiency in this Leigh's syndrome patient's cells was corrected by a nuclear
DNA-encoded factor from the HeLacot parent and ruled out an
mtDNA mutation as the basis for
COX deficiency. This system can be used to determine whether different generalized
mitochondrial disorders are due to mutations of nuclear or
mtDNA.