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Maltotriose consumption by hybrid Saccharomyces pastorianus is heterotic and results from regulatory cross-talk between parental sub-genomes

S. pastorianus strains are hybrids of S. cerevisiae and S. eubayanus that have been domesticated for several centuries in lager-beer brewing environments. As sequences and structures of S. pastorianus genomes are being resolved, molecular mechanisms and evolutionary origin of several industrially relevant phenotypes remain unknown. This study investigates how maltotriose metabolism, a key feature in brewing, may have arisen in early S. eubayanus × S. cerevisiae hybrids. To address this question, we generated a near-complete genome assembly of Himalayan S. eubayanus strains of the Holarctic subclade. This group of strains have been proposed to be the origin of the S. eubayanus subgenome of current S. pastorianus strains. The Himalayan S. eubayanus genomes harbored several copies of a SeAGT1 α-oligoglucoside transporter gene with high sequence identity to genes encountered in S. pastorianus. Although Himalayan S. eubayanus strains are unable to grown on maltose and maltotriose, their maltose-hydrolase and SeMALT1 and SeAGT1 maltose-transporter genes complemented the corresponding null mutants of S. cerevisiae. Expression, in a Himalayan S. eubayanus strain, of a functional S. cerevisiae maltose-metabolism regulator gene (MALx3) enabled growth on oligoglucosides. The hypothesis that the maltotriose-positive phenotype in S. pastorianus is a result of heterosis was experimentally tested by constructing a S. cerevisiae × S. eubayanus laboratory hybrid with a complement of maltose-metabolism genes that resembles that of current S. pastorianus strains. The ability of this hybrid to consume maltotriose in brewer’s wort demonstrated regulatory cross talk between sub-genomes and thereby validated this hypothesis. These results provide experimental evidence of the evolutionary origin of an essential phenotype of lager-brewing strains and valuable knowledge for industrial exploitation of laboratory-made S. pastorianus-like hybrids. Importance S.pastorianus, a S.cerevisiae X S.eubayanus hybrid, is used for production of lager beer, the most produced alcoholic beverage worldwide It emerged by spontaneous hybridization and have colonized early lager-brewing processes. Despite accumulation and analysis of genome sequencing data of S.pastorianus parental genomes, the genetic blueprint of industrially relevant phenotypes remains unknown. Assimilation of wort abundant sugar maltotriose has been postulated to be inherited from S.cerevisiae parent. Here, we demonstrate that although Asian S.eubayanus isolates harbor a functional maltotriose transporter SeAGT1 gene, they are unable to grow on α-oligoglucosides, but expression of S. cerevisae regulator ScMAL13 was sufficient to restore growth on trisaccharides. We hypothesized that S. pastorianus maltotriose phenotype results from regulatory interaction between S.cerevisae maltose transcription activator and the promoter of SeAGT1. We experimentally confirmed the heterotic nature of the phenotype and thus this results provide experimental evidence of the evolutionary origin of an essential phenotype of lager-brewing strains.