S101: Genome-wide screen of gene over-expression for ethanol resistance in Saccharomyces cerevisiae

Ethanol induces one of the most potent stresses faced by S. cerevisiae during industrial processes to produce this biofuel. Thus, engineering of resistance to ethanol is essential to improve yields and process efficiency.  Here we apply the Cytostat cell culture technique developed in our lab to screen for ethanol resistance phenotypes in S. cerevisiae cells transformed with a genome-wide, gene over-expression library. The main advantages of this approach are that, first, the Cytostat selects for inhibitor specific resistance because the continuous culture is maintained at very low cell densities, and thus the selective pressure is precisely defined by the composition of the feed stream.  This circumvents the possibility of resistance arising from the metabolic consumption or modification of the inhibitor, such that the resistance phenotype must be established by a sustainable mechanism in response to the constant challenge.  Most importantly, the Cytostat selects for the fittest, most resistant clone in the fastest possible way. Second, identifying the genetic modification only requires the sequencing of a small genomic fragment from the library and not of the whole genome. Using this approach, ethanol resistant clones, with over 50% improved growth rates under ethanol stress, were isolated from a pool of cells transformed with the library.  The effects of the over-expressed genes in the resistance phenotypes, including the kinetics of glucose consumption and ethanol production, cell size homeostasis, and gene expression will be presented.