Induced gene expression in industrial Saccharomyces pastorianus var. carlsbergensis TUM 34/70: evaluation of temperature and ethanol inducible native promoters

Induced gene expression is an important trait in yeast metabolic engineering, but current regulations prevent the use of conventional expression systems, such as galactose and copper, in food and beverage fermentations. This article examines the suitability of temperature-inducible native promoters for use in the industrial yeast strain Saccharomyces pastorianus var. carlsbergensis TUM 34/70 under brewing conditions. Ten different promoters were cloned and characterized under varying temperature shifts and ethanol concentrations using a green fluorescent protein reporter. The activities of these promoters varied depending upon the stress conditions applied. A temperature shift to 4 degrees C led to the highest fold changes of PSSA3, (P)UBI4 and (P)HSP104 by 5.4, 4.5 and 5.0, respectively. Ethanol shock at 24 degrees C showed marked, concentration-dependent induction of the promoters. Here, (P)HSP104 showed its highest induction at ethanol concentrations between 4% (v/v) and 6% (v/v). The highest fold changes of (P)SSA3 and (P)UBI4 were found at 10% (v/v) ethanol. In comparison, the ethanol shock at a typical fermentation temperature (12 degrees C) leads to lower induction patterns of these promoters. Taken together, the data show that three promoters ((P)HSP104, (P)UBI4 and (P)SSA3) have high potential for targeted gene expression in self-cloning brewing yeast using temperature shifts.