SUMMARY
Yeasts of Saccharomyces genus have long been associated with human beings. Their ability to convert different sugars into ethanol and CO2 has been used for making alcoholic beverages and food such as bread, setting the first example of  a biotechnological application of a microorganism. In recent years, yeast has played an important role as the most useful eukaryotic microorganism for biological analyses as well as for new developments in the field of Biotechnology.

The continued use for making alcoholic beverages and food of industrial strains of S.cerevisiae is justified by their ability to efficiently convert sugars into ethanol, carbon dioxide and many secondary metabolites that leads to balanced flavour and aroma in the final product and their ability to cope with stress caused mainly by temperature, osmotic pressure, hidrostatic pressure, high cellular density, ethanol and sharing with bacteria and other wild yeasts. However, we can improve stress tolerance achieving potential benefits in food and alcoholic beverages manufacturing processes. Fermentation at low temperature is proved to be a key factor in alcoholic beverages production with organoleptic characteristics that fit with sensory quality and consumer’s preference profiles.

Cellular response to downshift in temperature is not well characterized, and although is followed by the synthesis of a group of cold-inducible proteins, they are neither conserved nor shared by such a wide range of organisms as the heat shock proteins (HSPs).

The objective of the present Thesis is the identication and characterization of those genes that though overexpressión confer to S.cerevisiae strains a competitive adavantage to grow at low temperatures. Thus, we demonstrate that tryptophan prototrophy attenuates cold effects. In tryptophan auxotrophic strains, cold effects are mitigated with the addition of excess of tryptophan or with the overexpression of TRP1 (encodes phosphoribosylanthranilate isomerase that catalyzes the third step in tryptophan biosynthesis) or those genes that encode tryptophan transporters Tat2p and Tat1p. The overexpression of NSG2 (encodes a protein involved in regulation of sterol biosynthesis that stabilizes one of the two HMG-CoA isoenzymes), PCK1 (encodes phosphoenolpyruvate carboxykinase, key enzyme in gluconeogenesis) or PRO2 (encodes gamma-glutamyl phosphate reductase, catalyzes the second step in proline biosynthesis) cause an improvement in growth at 10º C in both tryptophan auxotrophic and tryptophan prototrophic strains, what means shorter lag phase and doubling time.

The overexpression in a prototrophic strain (RS-452)  of genes related to phosphate transport (PHO84, PHO87, PHO90 y GTR1) as well as the addition of inorganic phosphate, in order to attain a high phosphate concentration, improves its growth considerably.

Finally, we investigated the effects of low temperatures on tryptophan uptake through plasmatic membrane. At low temperatures tryptophan uptake is impaired both in laboratory strain W303 and the studied industrial strains. Individual overexpression of TAT2 and TAT1 causes a notable increase in tryptophan uptake at 10º C, which is correlated with an increased growth at that temperature.Taken together, these results suggest that tryptophan uptake is a key factor in yeast physiology, since under a variety of stress conditions it becomes a limiting factor for cell growth.