Proteome analysis of aerobically and anaerobically grown Saccharomyces cerevisiae cells.

The yeast Saccharomyces cerevisiae is able to grow under aerobic as well as anaerobic conditions. We and others previously found that transcription levels of approximately 500 genes differed more than two-fold when cells from anaerobic and aerobic conditions were compared. Here, we addressed the effect of anaerobic growth at the post-transcriptional level by comparing the proteomes of cells isolated from steady-state glucose-limited anaerobic and aerobic cultures. Following two-dimensional gel electrophoresis and mass spectrometry we identified 110 protein spots, corresponding to 75 unique proteins, of which the levels differed more than two-fold between aerobically and anaerobically-grown cells. For 21 of the 110 spots, the intensities decreased more than two-fold whereas the corresponding mRNA levels increased or did not change significantly under anaerobic conditions. The intensities of the other 89 spots changed in the same direction as the mRNA levels of the corresponding genes, although to different extents. For some genes of glycolysis a small increase in mRNA levels, 1.5-2 fold, corresponded to a 5-10 fold increase in protein levels. Extrapolation of our results suggests that transcriptional regulation is the major but not exclusive mechanism for adaptation of S. cerevisiae to anaerobic growth conditions.

Disruption of genes encoding pyruvate dehydrogenase kinases leads to retarded growth on acetate and ethanol in Saccharomyces cerevisiae.

Two open reading frames, YIL042c (PKP1) and YGL059w, with 25% sequence similarity to human pyruvate dehydrogenase kinases, were shown to have protein kinase activity. Using GFP fusions, it was demonstrated that the proteins localize in discrete submitochondrial regions. Strains with a null mutation in these loci grew poorly on acetate and ethanol as carbon sources. Doubling times increased from ca. 4 h in the wild-type to > 6 h for the mutants. Growth rates of the mutants could be restored to wild-type levels by simultaneous disruption of the PDA1 gene, encoding the E1alpha subunit of the pyruvate dehydrogenase complex. This observation and the pyruvate dehydrogenase activities measured in the mutant strains and the wild-type grown on glucose or acetate suggest that the slow growth phenotype on C2 carbon sources is caused by a futile cycle in which phosphoenolpyruvate is converted back to acetyl coenzyme A.