EXPRESSION AND FATE OF THE NUCLEARLY ENCODED SUBUNITS OF CYTOCHROME-C-OXIDASE IN CULTURED HUMAN-CELLS DEPLETED OF MITOCHONDRIAL GENE-PRODUCTS

Synthesis, import, assembly and turnover of the nuclearly encoded subu nits of cytochrome-e oxidase were investigated in cultured human cells depleted of mitochondrial gene products by continuous inhibition of m itochondrial protein synthesis (OP- cells). Immunoprecipitation after pulse labeling demonstrated that the synthesis of the nuclear subunits was not preferentially inhibited, implying that there is no tight reg ulation in the synthesis of mitochondrial and nuclear subunits of mito chondrial enzyme complexes. Quantitative analysis of the mitochondrial membrane potential in OP- cells indicated that its magnitude was abou t 30% of that in control cells. This explains the normal import of the nuclearly encoded subunits of cytochrome-e oxidase and other nuclearl y encoded mitochondrial proteins into the mitochondria that was found in OP- cells. The turnover rate of nuclear subunits of cytochrome-e ox idase, determined in pulse-chase experiments, showed a specific increa se in OP- cells. Moreover, immunoblotting demonstrated that the steady -state levels of nuclear subunits of cytochrome-e oxidase were severel y reduced in these cells, in contrast to those of the F-1 part of comp lex V. Native electrophoresis of mitochondrial enzyme complexes showed that assembly of the nuclear subunits of cytochrome-e oxidase did not occur in OP- cells, whereas the (nuclear) subunits of F-1 were assemb led. The increased turnover of the nuclear subunits of cytochrome-c ox idase in OP- cells is, therefore, most likely due to an increased susc eptibility of unassembled subunits to intra-mitochondrial degradation.