No abstract

The concentration of oxygen in the environment is one of the most important factors that regulate energy conversion in living cells. Organisms have developed multiple processes to optimize the utilization of oxygen when its availability is reduced. According to the role of oxygen in their metabolism, yeasts can be classified as: (a) obligate aerobes, displaying an exclusively respiratory metabolism; (b) facultative fermentatives, displaying both respiratory and fermentative metabolism; and (c) obligate fermentatives. The ability of yeasts to grow in very oxygen-limited conditions is strictly dependent on the ability to perform alcoholic fermentation, allowing reoxidation of NADH generated during glycolysis. In Saccharomyces cerevisiae, fermentation predominates over respiration when glucose concentrations are high, even under aerobic conditions. Depending on this characteristic, yeasts are classified as Crabtree-positive or Crabtreenegative. Thus, in Crabtree-positive yeasts, such as S. cerevisiae, NADH is mainly oxidized in glucose- The yeast Saccharomyces cerevisiae is characterized by its ability to: (a) degrade glucose or fructose to ethanol, even in the presence of oxygen (Crabtree effect); (b) grow in the absence of oxygen; and (c) generate respiratory-deficient mitochondrial mutants, so-called petites. How unique are these properties among yeasts in the Saccharomyces clade, and what is their origin? Recent progress in genome sequencing has elucidated the phylogenetic relationships among yeasts in the Saccharomyces complex, providing a framework for the understanding of the evolutionary history of several modern traits. In this study, we analyzed over 40 yeasts that reflect over 150 million years of evolutionary history for their ability to ferment, grow in the absence of oxygen, and generate petites. A great majority of isolates exhibited good fermentation ability, suggesting that this trait could already be an intrinsic property of the progenitor yeast. We found that lineages that underwent the whole-genome duplication, in general, exhibit a fermentative lifestyle, the Crabtree effect, and the ability to grow without oxygen, and can generate stable petite mutants. Some of the pre-genome duplication lineages also exhibit some of these traits, but a majority of the tested species are petite-negative, and show a reduced Crabtree effect and a reduced ability to grow in the absence of oxygen. It could be that the ability to accumulate ethanol in the presence of oxygen, a gradual independence from oxygen and ⁄ or the ability to generate petites were developed later in several lineages. However, these traits have been combined and developed to perfection only in the lineage that underwent the whole-genome duplication and led to the modern Saccharomyces cerevisiae yeast.Abbreviation EtBr, ethidium bromide.

When fruits ripen, microbial communities start a fierce competition for the freely available fruit sugars. Three yeast lineages, including baker’s yeast Saccharomyces cerevisiae, have independently developed the metabolic activity to convert simple sugars into ethanol even under fully aerobic conditions. This fermentation capacity, named Crabtree effect, reduces the cell-biomass production but provides in nature a tool to out-compete other microorganisms. Here, we analyzed over forty Saccharomycetaceae yeasts, covering over 200 million years of the evolutionary history, for their carbon metabolism. The experiments were done under strictly controlled and uniform conditions, which has not been done before. We show that the origin of Crabtree effect in Saccharomycetaceae predates the whole genome duplication and became a settled metabolic trait after the split of the S. cerevisiae and Kluyveromyces lineages, and coincided with the origin of modern fruit bearing plants. Our results suggest that ethanol fermentation evolved progressively, involving several successive molecular events that have gradually remodeled the yeast carbon metabolism. While some of the final evolutionary events, like gene duplications of glucose transporters and glycolytic enzymes, have been deduced, the earliest molecular events initiating Crabtree effect are still to be determined.

The origin of modern fruits brought to microbial communities an abundant source of rich food based on simple sugars. Yeasts, especially Saccharomyces cerevisiae, usually become the predominant group in these niches. One of the most prominent and unique features and likely a winning trait of these yeasts is their ability to rapidly convert sugars to ethanol at both anaerobic and aerobic conditions. Why, when, and how did yeasts remodel their carbon metabolism to be able to accumulate ethanol under aerobic conditions and at the expense of decreasing biomass production? We hereby review the recent data on the carbon metabolism in Saccharomycetaceae species and attempt to reconstruct the ancient environment, which could promote the evolution of alcoholic fermentation. We speculate that the first step toward the so-called fermentative lifestyle was the exploration of anaerobic niches resulting in an increased metabolic capacity to degrade sugar to ethanol. The strengthened glycolytic flow had in parallel a beneficial effect on the microbial competition outcome and later evolved as a “new” tool promoting the yeast competition ability under aerobic conditions. The basic aerobic alcoholic fermentation ability was subsequently “upgraded” in several lineages by evolving additional regulatory steps, such as glucose repression in the S. cerevisiae clade, to achieve a more precise metabolic control.

Saccharomyces yeasts degrade sugars to two-carbon components, in particular ethanol, even in the presence of excess oxygen. This characteristic is called the Crabtree effect and is the background for the 'make–accumulate–consume' life strategy, which in natural habitats helps Saccharomyces yeasts to out-compete other microorganisms. A global promoter rewiring in the Saccharomyces cerevisiae lineage, which occurred around 100 mya, was one of the main molecular events providing the background for evolution of this strategy. Here we show that the Dekkera bruxellensis lineage, which separated from the Saccharomyces yeasts more than 200 mya, also efficiently makes, accumulates and consumes ethanol and acetic acid. Analysis of promoter sequences indicates that both lineages independently underwent a massive loss of a specific cis-regulatory element from dozens of genes associated with respiration, and we show that also in D. bruxellensis this promoter rewiring contributes to the observed Crabtree effect.