Sulphate Uptake Plays a Major Role in the Production of Sulphur Dioxide by Yeast Cells during Oenological Fermentations

Sulphur dioxide (SO2) is mostly used as an antioxidant additive in winemaking, but excessive levels may be harmful to both wine quality and consumers health. During fermentation, yeast Saccharomyces cerevisiae contributes significantly to final SO2 levels, and low-producing strains become especially interesting for the wine industry. Recent evidence implicating the impairment of sulphate transport in the SO2 decrease prompted us to further investigate the sulphate/sulphite metabolic connection in multiple winery yeast strains. Here, we inactivated by CRISPR/Cas9 the high-affinity sulphate permeases (Sul1p and Sul2p) in four strains normally used in winemaking, selected by their different abilities to produce SO2. Mutant strains were then used to perform fermentation assays in different types of natural must, and the final levels of SO2 and other secondary metabolites, crucial for wine organoleptic properties, were further determined for all fermentation products. Overall, data demonstrated the double Delta SUL1/Delta SUL2 inactivation in winery strains significantly decreases the levels of SO2 produced by mutant cells, without however altering both yeast fermentative properties and the ability to release relevant metabolites. Since similar effects were observed in diverse must types for strains with different features, the data strongly support that sulphate assimilation is the determining factor in SO2 production during oenological fermentations.