Abstract
Analyses of cellular processes in the yeast Saccharomyces cerevisiae rely primarily upon a small number of highly domesticated laboratory strains, leaving the extensive natural genetic diversity of the model organism largely unexplored and unexploited. We asked if this diversity could be used to enrich our understanding of basic biological processes. As a test case, we examined a simple trait: the utilization of di/tripeptides as nitrogen sources. The capacity to import small peptides is likely to be under opposing selective pressures (nutrient utilization versus toxin vulnerability) and may therefore be sculpted by diverse pathways and strategies. Hitherto, dipeptide utilization in S. cerevisiae was solely ascribed to the activity of a single protein, the Ptr2p transporter. Using high-throughput phenotyping and several genetically diverse strains, we identified previously unknown cellular activities that contribute to this trait. We find that the Dal5p allantoate/ureidosuccinate permease is also capable of facilitating di/tripeptide transport. Moreover, even in the absence of Dal5p and Ptr2p, an additional activity—almost certainly the periplasmic asparaginase II Asp3p—facilitates the utilization of dipeptides with C-terminal asparagine residues by a different strategy. Another, as-yet-unidentified activity enables the utilization of dipeptides with C-terminal arginine residues. The relative contributions of these activities to the utilization of di/tripeptides vary among the strains analyzed, as does the vulnerability of these strains to a toxic dipeptide. Only by sampling the genetic diversity of multiple strains were we able to uncover several previously unrecognized layers of complexity in this metabolic pathway. High-throughput phenotyping facilitates the rapid exploration of the molecular basis of biological complexity, allowing for future detailed investigation of the selective pressures that drive microbial evolution.
Highlights
Our understanding of the inner workings of eukaryotic cells owes much to the yeast Saccharomyces cerevisiae
To advance our understanding of the processes contributing to nitrogen utilization in S. cerevisiae, we studied a panel of strains isolated from several widely differing growth environments (Table 1)
Because expression of the dipeptide transporter PTR2 is strongly induced by even micromolar amounts of certain amino acids supplemented in the media [29], we first restored all of these strains to full amino acid prototrophy
Summary
Our understanding of the inner workings of eukaryotic cells owes much to the yeast Saccharomyces cerevisiae. The application of powerful genetic and molecular tools to this model organism has yielded an extensively annotated proteome These analyses have benefited greatly from the engineering of experimentally tractable strains of S. cerevisiae, but an unintended consequence of this focus has been a tendency to ignore the vast wealth of natural genetic variation found in diverse strains of this organism. Phenotypic analyses of diverse yeast strains [1,2] and the application of microarray technology to the analysis of allelic variation [3,4,5,6,7,8] and population genetic variation in gene expression [9,10,11] are providing new insights into the ecology and diversity of the species. We employed highthroughput phenotyping of diverse S. cerevisiae strains to dissect the multiple activities contributing to the utilization of di/tripeptides as a nitrogen source
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