[9] The Isolation of Mitochondrial and Nuclear Mutants of Saccharomyces cerevisiae with Specific Defects in Mitochondrial Functions

This chapter reviews the biogenesis of respiratory competent mitochondria in the yeast Saccharomyces cerevisiae that is dependent on the function of both mitochondrial and nuclear genes. Mitochondrial DNA includes genes that determine and/or regulate the biosynthesis of subunit polypeptides of the respiratory enzymes cytochrome oxidase and coenzyme QH₂-cytochrome C reductase and of the oligomycin-sensitive ATPase. It is known that mitochondrial DNA contains genes that code for the ribosomal RNA's and transfer RNA's of the mitochondrial protein synthetic machinery. These are known as “syn genes.” The chapter discusses that the contribution of the nuclear DNA to the respiratory competence of mitochondria includes structural and regulatory genes for the matrix enzymes, the DNA replication and transcription systems, and the proteins of the ribosomes, the F₁ ATPase, and certain subunits of the respiratory enzyme complexes. Strains that carry nuclear mutations resulting in an inability to use nonfermentable substrates for growth are called “pet mutants.” Mutant isolation procedures described in the chapter are designed to select mutants with specific defects in mitochondrial functions. They include (1) pet mutants with specific lesions in the respiratory enzymes or the ATPase, (2) mit^– mutants with lesions in cytochrome oxidase, coenzyme QH₂-cytochrome C reductase or oligomycin-sensitive ATPase, and (3) syn– mutants specifically defective in mitochondrial protein synthesis.