Surf One 74

ABSTRACT

C173 and W125 are pet mutants of Saccharomyces cerevisiae, partially deficient in cytochrome oxidase but with elevated concentrations of cytochrome c. Assays of electron transport chain enzymes indicate that the mutations exert different effects on the terminal respiratory pathway, including an inefficient transfer of electrons between the bc1 and the cytochrome oxidase complexes. A cloned gene capable of restoring respiration in C173/U1 and W125 is identical to reading frame YGR112w of yeast chromosome VII (GenBank Z72897[GenBank]). The encoded protein is homologous to the product of the mammalian SURF-1 gene. In view of the homology, the yeast gene has been designated SHY1 (Surf Homolog of Yeast). An antibody against the carboxyl-terminal half of Shy1p has been used to localize the protein in the inner mitochondrial membrane. Deletion of part of SHY1 produces a phenotype similar to that of G91 mutants. Disruption of SHY1 at a BamHI site, located approximately 2/3 of the way into the gene, has no obvious phenotypic consequence. This evidence, together with the ability of a carboxyl-terminal coding sequence starting from the BamHI site to complement a shy1 mutant, suggests that the Shy1p contains two domains that can be separately expressed to form a functional protein.

INTRODUCTION

Respiratory defective pet mutants1 of Saccharomyces cerevisiae have been useful in understanding some of the processes underlying the biogenesis of mitochondria. In earlier studies, pet mutants have been grouped into different phenotypic classes based on their spectral properties and their respiratory and ATPase activities (1, 2). A substantial number of complementation groups consist of mutants displaying defects in single enzymes of the respiratory chain (e.g. cytochrome oxidase, ubiquinol-cytochrome c reductase) or ATPase. Such strains have been exploited in different laboratories to identify and isolate genes coding for the protein constituents of the respiratory complexes (3, 4) and for ancillary factors acting at different stages of their assembly (5-7). Another commonly encountered phenotype is characterized by the pleiotropic absence of oligomycin-sensitive ATPase and the respiratory chain complexes, whose synthesis depends in part on the expression of the mitochondrial gene products. This phenotype is frequently elicited by mutations in constituents of the mitochondrial translational machinery (8).

In addition, numerous mutants exhibit the presence of all the respiratory chain components but at levels below those found in wild type yeast. This class of mutants has received little attention because the gross biochemical phenotypes do not provide obvious clues about the primary lesions responsible for the respiratory defect. To enlarge on current information about the contribution of the nuclear genome toward the maintenance of respiratory competent mitochondria, we have begun studies of pet mutants with partial pleiotropic phenotypes. In this communication we report on mutants from complementation group G91 of ojavascript:void(0)ur collection of pet strains (9). Enzyme assays have revealed that the depression of mitochondrial respiration in this group of mutants cannot be explained in any simple way by the activities of different segments of the electron transport chain but rather appears to be related to inefficient transfer of electrons in the span between the bc1 and cytochrome oxidase complexes. The mutations responsible for the respiratory defect have been localized to the yeast homolog of the SURF-1 gene, previously shown to be in a highly conserved gene cluster of several mammalian (10) and chicken genomes (11). We present evidence that the product of the yeast SHY1 gene is a membrane constituent of mitochondria and therefore is likely to function directly in some aspect of mitochondrial organization and function.