Critical role of RPI1 in the stress tolerance of yeast during ethanolic fermentation.

Stress tolerance of yeast Saccharomyces cerevisiae during ethanolic fermentation is poorly understood due to the lack of genetic screens and conventional plate assays for studying this phenotype. We screened a genomic expression library of yeast to identify gene(s) that, upon overexpression, would prolong the survival of yeast cells during fermentation, with the view to understand the stress response better and to use the identified gene(s) in strain improvement. The yeast RPI1 (Ras-cAMP pathway inhibitor 1) gene was identified in such a screen performed at 38 degrees C; introducing an additional copy of RPI1 with its native promoter helped the cells to retain their viability by over 50-fold better than the wild type (WT) parent strain, after 36 h of fermentation at 38 degrees C. Disruption of RPI1 resulted in a drastic reduction in viability during fermentation, but not during normal growth, further confirming the role of this gene in fermentation stress tolerance. This gene seems to improve viability by fortifying the yeast cell wall, because RPI1 overexpression strain is highly resistant to cell lytic enzyme zymolyase, compared with the WT strain. As the RPI1 overexpression strain substantially retains cell viability at the end of fermentation, the cells can be reused in the subsequent round of fermentation, which is likely to facilitate economical production of ethanol.