The physical and genetic map of the 73 kb mitochondrial genome of a novel yeast species Saccharomyces douglasii was determined. All known mitochondrial genes were localised: the 3 mitochondrially encoded subunits of cytochrome c oxydase (COXI, COXII and COXIII); the 3 subunits of adenosine triphosphatase (ATPase) complex (subunits 6, 8 and 9); apocytochrome b; the ribosomal protein VAR 1; the 2 ribosomal ribonucleic acid (RNA); the transfer RNA (tRNA) maturation locus TSL and all tRNAs. All these genes were sequenced and compared with their counterparts from S. cerevisiae.
Molecular analysis of mitochondrial deoxyribonucleic acid (mtDNA) from a respiratory sufficient recombinant (issued from an interspecific cross between parents which did not respire) has shown that restoration of respiration is directly correlated with the absence of the first intron of S. douglasii COXI gene.
The S. douglasii nuclear gene able to promote the excision of the first intron of mitochondrialCOXI gene is the homologue of the S. cerevisiae MRS 1 gene, which is essential for the excision of the 2 mitochondrial introns (bi3 of cytochrome b gene and ai5beta of COXI gene), but is unable to assure the excision of the first intron from the COXI gene of S. douglasii.
The CBP2 gene in S. cerevisiae is responsible for the excision of the last intron of CYTb gene, which is lacking in S. douglasii gene. However, the S. douglasii CBP2 gene promotes the splicing of the S. douglasii intron located in LSrRNA gene. The divergence of the MRS1 and CBP2 genes of S. cerevisiae and S. douglasii, and concomitant changes in the structure of their mitochondrial genomes is an interesting example of the coevolution of nuclear and mitochondrial genomes.
The mitochondrial exonic sequences are remarkably conserved (only 2% of substitutions) between the 2 species in strong contrast with relatively important divergence observed (12%) for their nuclear genes. Interestingly, this is the reverse of the situation observed in the vertebrate system, in which mitochondrial genes accumulate 5 to 10-fold more nucleotide substitutions than the nuclear ones.
All eucaryotic cells contain two separate genomes : the nuclear & the mitochondrial. A novel approach to the study of nucleo-mitochondrial interactions will be used. It is based on the analysis of interspecific hybrids which combine the nuclear genome of one species with the mitochondiral genome of another species and display a nucleo-mitochondrial incompatibility. The structural, recombinationational and functional analyses will provide new informations in terms of molecular mechamisms involved and evolutionary comparisons on protein and RNA coding regions, introns and regulatory signals in the nuclear and mtDNA's. Novel mitochondrial genomes will be constructed and the structure and properties of nuclear genes controlling their expression will be analysed.
Funding SchemeCSC - Cost-sharing contracts