Effect of TDA1 deletion on cell cycle state in S. cerevisiae during nutrient limitation

Protein kinases are enzymes which phosphorylate other proteins and often play critical regulatory roles in cellular metabolism. We studied the effects of deletion of the TDA1 gene in Saccharomyces cerevisiae. TDA1 encodes a protein kinase of largely unknown function in yeast, but its human ortholog, NUAK1, has been implicated in oncogenesis. We initially examined the growth rate or the mean rate of increase in size of tda1 mutants. These preliminary studies showed that homozygous diploid tda1 mutants have an approximately 15-20% reduced growth rate in galactose and glycerol media, compared to the wild-type. These results suggest loss of TDA1 leads to a nutrient-specific metabolic deficiency. In order to elucidate the effect of the tda1 deletion on metabolism, asynchronous yeast cultures were grown in chemostats in glycerol and ammonium nutrient-limited media. Individual cells from both tda1Δ and WT samples taken at different dilution rates were analyzed for budding index and DNA content using flow cytometry. This analysis examines the cell cycle state of a yeast population under specific nutrient limitation at a given dilution rate. Glycerol and nitrogen limitations were tested. We found that the mutant seems to have a reduced ability to produce glycerol. These experiments provide valuable data on the metabolic function of Tda1p and its role in the relationship between cell division and metabolism