Isolation of a nuclear yeast gene involved in the mitochondrial import of cytoplasmically synthesized precursor proteins

Abstract

Several nuclear mutants of the yeast, Saccharomyces cerevisiae, have been characterized which synthesize only the higher-molecular weight precursor but not the mature subunit VI of the mitochondrial ubiquinol cytochrome c oxidoreductase. The mutants belong to different complementation groups and vary in the extent of their being simultaneously deficient in other components of the mitochondrial inner membrane. From a yeast genomic DNA library the plasmid pTS2326 was isolated which complements the defect in one of these mutants, ts2326. The cloned DNA fragment, 2.3 kilobases in length, was sequenced. It contains two open reading frames, ORF1 and ORF2, consisting of 723 and 417 base pairs, respectively. By selective deletion of either reading frame it was shown that only ORF1 containes the information necessary to complement the ts2326 mutation. The ORF1 coding sequence is not the structural gene of subunit VI. The postulated gene product of ORF1 has a molecular weight of 27.114 daltons. It exhibits several sequence characteristics typical of proteins which are internalized by the mitochondrial membrane systems. It is proposed that ORF1 is involved in the import and processing of cytoplasmically synthesized mitochondrial precursors.