Lipid composition of subcellular membranes of an FY1679-derived haploid yeast wild-type strain grown on different carbon sources.

The aim of the project EUROFAN (European Functional Analysis Network) is to elucidate the function of unknown genes of the yeast Saccharomyces cerevisiae at a large scale. Functional analysis is based on general and specific tests with yeast deletion strains. A prerequisite for these studies is a profound knowledge of the biochemistry and cell biology of the corresponding wild-type strain FY1679.

Deletion of six open reading frames from the left arm of chromosome IV of Saccharomyces cerevisiae.

The construction of six deletion mutants of Saccharomyces cerevisiae and their basic phenotypic characterization are described. Open reading frames YDL148c, YDL109c, YDL021w, YDL019c, YDL018c and YDL015c from the left arm of chromosome IV were deleted using a polymerase chain reaction (PCR)-based disruption technique, introducing the kanMX4 resistance marker into the respective genes. Gene replacement cassettes (pYORCs) for use in other strain backgrounds were cloned by PCR using DNA templates from haploid or diploid deletion mutants, and inserted into episomal plasmids.

Systematic analysis of yeast strains with possible defects in lipid metabolism.

Lipids are essential components of all living cells because they are obligate components of biological membranes, and serve as energy reserves and second messengers. Many but not all genes encoding enzymes involved in fatty acid, phospholipid, sterol or sphingolipid biosynthesis of the yeast Saccharomyces cerevisiae have been cloned and gene products have been functionally characterized. Less information is available about genes and gene products governing the transport of lipids between organelles and within membranes or the turnover and degradation of complex lipids.

YDL142c encodes cardiolipin synthase (Cls1p) and is non-essential for aerobic growth of Saccharomyces cerevisiae.

The unassigned open reading frame YDL142c was identified to code for cardiolipin synthase, Cls1p. A cls1 deletion strain is viable on glucose, galactose, ethanol, glycerol and lactate containing media, although the growth rate on non-fermentable carbon sources is decreased. Mitochondria of the cls1 mutant are devoid of cardiolipin but accumulate the cardiolipin precursor phosphatidylglycerol when grown on non-fermentable carbon sources.

Mrs5p, an essential protein of the mitochondrial intermembrane space, affects protein import into yeast mitochondria.

We have isolated a yeast nuclear gene that suppresses the previously described respiration-deficient mrs2-1 mutation when present on a multicopy plasmid. Elevated gene dosage of this new gene, termed MRS5, suppresses also the pet phenotype of a mitochondrial splicing-deficient group II intron mutation M1301. The MRS5 gene product, a 13-kDa protein of low abundance, shows no similarity to other known proteins and is associated with the inner mitochondrial membrane, protruding into the intermembrane space.