In Saccharomyces cerevisiae, the inner mitochondrial membrane protein complex cytochrome c oxidase is composed of nine non-identical polypeptide subunits. The three largest subunits (I-III) are encoded in the mitochondrial genome, while the structural genes for the remaining six subunits (IV-VII, VIIa and VIII) reside in the nucleus. One of these subunits, subunit V, is unique in several respects. First, it is apparently the only subunit encoded by multiple genes. Second, the two subunit V genes, COX5a and COX5b are not alike. They share only 66% amino acid sequence homology, although either polypeptide is fully capable of functioning as a bona fide subunit of the holoenzyme. Third, while it is the COX5a gene product that is usually found in mitochondria, transcripts for both genes are present in approximately equal amounts under most conditions, suggesting that COX5b is regulated at a level other than transcription. Finally, while the COX5a gene is continuous, the COX5b gene contains an intron that appears to be inefficiently processed. An alteration in a highly conserved splice sequence, (a GCATGT instead of a GTATGT at the 5' splice junction), may account, in part, for the splicing inadequacy, and the regulation of this gene. We propose to study the biology and regulation of this small gene family, and to understand molecularly what factor(s) are responsible for the presence of either subunit, Va or Vb, within yeast mitochondria and holo-cytochrome c oxidase. In order to achieve this goal, we plan to determine in detail how these genes are regulated and how they respond respond to different environmental stimuli. These studies will address regulation at the levels of transcription, translation, and protein turnover, and will also examine the role of RNA processing in the regulation of COX5b. We will identify sequences that act in cis, to influence the expression of these genes, as well as determine if the product(s) of unlinked genes act in trans to regulate COX5a and COX5b. This system, within the powerful experimental background of yeast, provides an opportunity to study the differential expression of two genes within a gene family. A such, it should provide insights towards our understanding of the biology and biogenesis of an important inner membrane protien complex, and of mitochondria in general.
