Abstract
The lager-brewing yeast Saccharomyces pastorianus is a hybrid between S. cerevisiae and S. eubayanus with an exceptional degree of aneuploidy. While chromosome copy number variation (CCNV) is present in many industrial Saccharomyces strains and has been linked to various industrially-relevant traits, its impact on the brewing performance of S. pastorianus remains elusive. Here we attempt to delete single copies of chromosomes which are relevant for the production of off-flavor compound diacetyl by centromere silencing. However, the engineered strains display CNV of multiple non-targeted chromosomes. We attribute this unintended CCNV to inherent instability and to a mutagenic effect of electroporation and of centromere-silencing. Regardless, the resulting strains displayed large phenotypic diversity. By growing centromere-silenced cells in repeated sequential batches in medium containing 10% ethanol, mutants with increased ethanol tolerance were obtained. By using CCNV mutagenesis by exposure to the mitotic inhibitor MBC, selection in the same set-up yielded even more tolerant mutants that would not classify as genetically modified organisms. These results show that CCNV of alloaneuploid S. pastorianus genomes is highly unstable, and that CCNV mutagenesis can generate broad diversity. Coupled to effective selection or screening, CCNV mutagenesis presents a potent tool for strain improvement.
Highlights
The lager brewing yeast Saccharomyces pastorianus is an interspecific hybrid of Saccharomyces cerevisiae and the cold-tolerant Saccharomyces eubayanus (Libkind et al, 2011; Salazar et al, 2019)
In order to assess the phenotypic impact of chromosome CNV in an alloaneuploid S. pastorianus genome, we attempted to delete copies of individual chromosomes in strain CBS 1483 (Bolat et al, 2013; Brickwedde et al, 2017)
Saccharomyces pastorianus is an interspecific hybrid of S. cerevisiae and S. eubayanus (Nakao et al, 2009; Libkind et al, 2011) that has been domesticated in Europe since the late Middle Ages (Meussdoerffer, 2009) and that accounts for 89% of brewed beer worldwide
Summary
The lager brewing yeast Saccharomyces pastorianus is an interspecific hybrid of Saccharomyces cerevisiae and the cold-tolerant Saccharomyces eubayanus (Libkind et al, 2011; Salazar et al, 2019) Chromosomes from both parental species are present in the genome of S. pastorianus in varying number of copies, making the genome alloploid and aneuploid (Dunn and Sherlock, 2008). Engineering Yeast Chromosome Copy Number an approximately diploid S. eubayanus subgenome, Group 1 strains (Saaz) have a, generally incomplete, haploid S. cerevisiae subgenome, while Group 2 strains (Frohberg) have a diploid or higher S. cerevisiae subgenome (Nakao et al, 2009; van den Broek et al, 2015; Okuno et al, 2016) Reflecting these differences in genome composition, Group 1 strains are more cold-tolerant whereas Group 2 strains exhibit more efficient maltotriose consumption, traits associated with S. eubayanus and S. cerevisiae, respectively (Gibson et al, 2013; Brouwers et al, 2019a,b). There are large copy number differences between the genomes of S. pastorianus strains, even among supposedly clonal isolates, suggesting high genomic plasticity and chromosome copy number instability (Bolat et al, 2008; van den Broek et al, 2015)
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
