Date Published: February 12, 2018
Publisher: Public Library of Science
Author(s): Claire Brice, Francisco A. Cubillos, Sylvie Dequin, Carole Camarasa, Claudio Martínez, Joseph Schacherer.
Saccharomyces cerevisiae strains are genetically diverse, largely as a result of human efforts to develop strains specifically adapted to various fermentation processes. These adaptive pressures from various ecological niches have generated behavioral differences among these strains, particularly in terms of their nitrogen consumption capacities. In this work, we characterize this phenotype by the specific quantity of nitrogen consumed under oenological fermentation conditions using a new approach. Indeed, unlike previous studies, our experiments were conducted in an environment containing excess nitrogen, eliminating the nitrogen limitation/starvation factor that is generally observed in fermentation processes. Using these conditions, we evaluated differences in the nitrogen consumption capacities for a set of five strains from diverse origins. The strains presented extremely different phenotypes and variations in their capacities to take up nitrogen from a wine fermentation environment. These variations reflect the differences in the nitrogen uptake capacities between wine and non-wine strains. Finally, the strains differed in their ability to adapt to the nitrogen composition of the environment, leading to variations in the cellular stress states, fermentation performances and the activity of the nitrogen sensing signaling pathway.
The budding yeast Saccharomyces cerevisiae is the most exploited microorganism in the food industry because of its ability to achieve complete fermentation of solutions with high sugar contents, and the sugars are converted into alcohol, carbon dioxide and secondary end-products. Different studies have demonstrated the wide genetic diversity of S. cerevisiae strains [1–4] that result from the combination of their natural genetic diversity (or non-domesticated yeasts) and different domestication processes. Yeasts occupy diverse natural habitats, such as rotten fruits, flowering plant nectars, hops and tree exudates [5–6]. The selective pressure imposed by these stressful environmental conditions has clearly impacted the evolution of these species. Furthermore, human activity has shaped the genetics of the yeast population to obtain yeasts with adaptive properties for use in several industrial fermentation processes, such as baking, brewing, winemaking and the production of various fermented beverages [7–9]. Consequently, S. cerevisiae strains can be classified in five distinct lineages according to their geographic origin or isolation sources (Saccharomyces Genome Resequencing Project (SGRP)); , some of them (wine and sake lineages) are the result of distinct domestication events .
Phenotypic diversity among yeast strains has long been characterized and exploited, particularly with regards to the central carbon metabolism . However, some aspects of the differences in the abilities of strains to efficiently metabolize nitrogen compounds remain unclear. Previous studies have demonstrated the substantial diversity of nitrogen consumption capacities within S. cerevisiae species [26, 38, 43, 45], which is partly related to their original environment. It has also been reported that the differences between strains can be amplified by providing excess nitrogen . In this study, the comparison of the nitrogen consumption profiles of five S. cerevisiae strains, originating from nitrogen-poor (NA, WA and SA) or nitrogen-rich (WE, FWI) environments, confirmed these differential behaviors. The WE and FWI strains consumed up to 20% more YAN during fermentations with excess nitrogen than the NA, WA and SA strains. Furthermore, we demonstrated that the differences between the capacities of the strains to consume YAN were a result of variations in their abilities to uptake specific nitrogen sources, and the residual amounts of these sources at the end of the growth phase varied between the WE and FWI strains on one hand and the NA, WA and SA on the other hand. The differentially transported N compounds included branched amino acids (leucine, valine and isoleucine) transported by the permeases Bap2p and Bap3p , the aromatic amino acids (tyrosine, phenylalanine) transported by the permeases Tat1p and Tat2p , glutamic acid transported by the permease Dip5p  and ammonium ions exclusively transported by the Mep1p, Mep2p and Mep3p permeases [27–28]. The variations the capacities of these strains to import these nitrogen sources may be due to mutations in the coding sequences that modulate the activities of the permeases or to differences in the expression pattern of genes encoding for these transporters. Comparison of the coding sequences and reciprocal hemizygosity analysis for the candidate genes exhibiting non-synonymous mutations allowed us to rule out the contribution of these mutations to the differences in the abilities of the strains to import nitrogen compounds.
In this study, we investigated the metabolic and molecular basis for the differences in nitrogen consumption between strains. Using strains exhibiting extremely diverse nitrogen consumption profiles, we characterized this phenotype based on the variability between strains in their abilities to consume different quantities of YAN during the fermentation process. We determinate that variations in nitrogen consumption capacity between strains are associated with different adaptation capacities of the strains for the winemaking environment. Thus, the efficiency of nitrogen consumption seems to be involved in the ability of the strains to adapt to the winemaking environment. It would be interesting to know if variations in nitrogen consumption capacity reflect differences in the nitrogen metabolic requirements between the wine and non-wine strains. Further investigations are now in progress to identify the cause of this variability by exploring the management of the nitrogen anabolic requirements between these strains.