Identification of the fitness determinants of budding yeast on a natural substrate

Abstract

The budding yeasts are prime models in genomics and cell biology, but the ecological factors that determine their success in non-human-associated habitats is poorly understood. In North America Saccharomyces yeasts are present on the bark of deciduous trees, where they feed on bark and sap exudates. In the North East, Saccharomyces paradoxus is found on maples, which makes maple sap a natural substrate for this species. We measured growth rates of S. paradoxus natural isolates on maple sap and found variation along a geographical gradient not explained by the inherent variation observed under optimal laboratory conditions. We used a functional genomic screen to reveal the ecologically relevant genes and conditions required for optimal growth in this substrate. We found that the allantoin degradation pathway is required for optimal growth in maple sap, in particular genes necessary for allantoate utilization, which we demonstrate is the major nitrogen source available to yeast in this environment. Growth with allantoin or allantoate as the sole nitrogen source recapitulated the variation in growth rates in maple sap among strains. We also show that two lineages of S. paradoxus display different life-history traits on allantoin and allantoate media, highlighting the ecological relevance of this pathway.

Figure 1

Wild strains of Saccharomyces show variation in growth parameters on maple sap. (a) Principal component analysis on correlations of growth rates and efficiency measured for 73 strains on six maple saps. PC1 and PC2 positively reflect growth efficiency and rate in maple sap samples, respectively. Colored symbols correspond to S. paradoxus lineages as defined in Leducq et al. (2016): SpA (green squares), SpB (red circles), SpC (blue triangles), SpC* (purple diamonds), SpBf (orange rectangle) and the empty square represents a wild S. cerevisiae strain. (b) Map showing the relationship between the median PC2 and the geographical origin of the strains. The area defined by the broken line represents the distribution of Acer saccharum Marsh (based on USGS.gov).

Figure 2

The most relevant genes for growth in maple samples are enriched in biological processes relevant for nitrogen utilization. (a) The lists of genes with significant effects were defined based on the median fitness difference with the control and the median P-value of a Welch test for maple saps and syrups. (b) Functionally grouped network with biological processes as nodes linked based on their κ-score level (⩾ 0.4). Node color represents the term enrichment specificity to sap, syrup or both gene lists and the node size reflects the significance.

Figure 3

The S. cerevisiae deletion strains for genes involved in the allantoin catabolic pathway and its regulation show consistent fitness differences in maple samples relative to the control media. (a) Members of the allantoin catabolic pathway and its known regulators, including genes regulating the nitrogen catabolic repression, found among the candidates are highlighted in bold. Genes are colored according to their overall median fitness difference from the control. (b) The detailed fitness effect of each gene deletion in the maple samples over the flow period connected by dotted lines (DAL pathway) or full lines (Regulators). The YKR033C ORF overlaps the DAL80 gene and was among the candidates.

Figure 4

Quantification of allantoin and allantoate in maple sap concentrates (a) and maple syrups (b) echoes growth efficiency on maple sap and syrup samples over the sampling flow period (c). Box plots represent the distribution of growth efficiency of 73 wild Saccharomyces strains in each sample. Whiskers indicate the 1.5 interquartile range.

Figure 5

Heat map showing the growth efficiency of S. paradoxus knockout strains of the allantoin catabolism pathway compared with the wild type (WT) on maple samples and synthetic media with allantoate (ALA), allantoin (ALN) or ammonium sulfate (AMS) as the nitrogen source. DAL2 and DUR1,2 are required for growth on most maple saps while DAL1 and DAL7 are not. These results indicate that deletion strains unable to utilize allantoate as a nitrogen source are unable to grow on most maple sap samples, underlying the absence of alternate nitrogen sources.

Figure 6

Variation in growth rate among strains in maple saps does not correlate with growth rate in rich undefined media (YPD). Minimal media containing sucrose and ammonium sulfate (AMS) show intermediate correlations, which can be increased by using allantoate (ALA) or allantoin (ALN) as the nitrogen source. Dots are colored according to the correlation coefficient.

Figure 7

Correlations between growth parameters of S. paradoxus lineages SpB and SpC in maple saps (a) and in rich undefined (YPD) and synthetic media containing either ammonium sulfate (AMS), allantoin (ALN) or allantoate (ALA) (b).