An Evaluation of High-Throughput Approaches to QTL Mapping in <i>Saccharomyces cerevisiae</i>

Wilkening, Stefan; Lin, Gen; Fritsch, E.; Tekkedil, Manu M.; Anders, Simon; Kuehn, Raquel; Nguyen, Michelle; Aiyar, Raeka S.; Proctor, Michael; Sakhanenko, Nikita A.; Galas, David J.; Gagneur, Julien; Deutschbauer, Adam M.; Steinmetz, Lars M.

doi:10.1534/genetics.113.160291

Cited by 79 publications

(94 citation statements)

References 77 publications

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“…Transformation is known to be mutagenic (Shortle et al 1984), and mutations can accumulate during strain construction (Wilkening et al 2014). We advise that future studies carefully control for plasmid copy number.…”

Section: Discussionmentioning

confidence: 99%

Comprehensive Analysis of the SUL1 Promoter of Saccharomyces cerevisiae

et al. 2016

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In the yeast Saccharomyces cerevisiae, beneficial mutations selected during sulfate-limited growth are typically amplifications of the SUL1 gene, which encodes the high-affinity sulfate transporter, resulting in fitness increases of .35% . Cis-regulatory mutations have not been observed at this locus; however, it is not clear whether this absence is due to a low mutation rate such that these mutations do not arise, or they arise but have limited fitness effects relative to those of amplification. To address this question directly, we assayed the fitness effects of nearly all possible point mutations in a 493-base segment of the gene's promoter through mutagenesis and selection. While most mutations were either neutral or detrimental during sulfate-limited growth, eight mutations increased fitness .5% and as much as 9.4%. Combinations of these beneficial mutations increased fitness only up to 11%. Thus, in the case of SUL1, promoter mutations could not induce a fitness increase similar to that of gene amplification. Using these data, we identified functionally important regions of the SUL1 promoter and analyzed three sites that correspond to potential binding sites for the transcription factors Met32 and Cbf1. Mutations that create new Met32-or Cbf1-binding sites also increased fitness. Some mutations in the untranslated region of the SUL1 transcript decreased fitness, likely due to the formation of inhibitory upstream open reading frames. Our methodology-saturation mutagenesis, chemostat selection, and DNA sequencing to track variants-should be a broadly applicable approach.KEYWORDS high-throughput sequencing; transcription; promoter mutagenesis; gene amplification; chemostat; yeast; Saccharomyces cerevisiae C HANGES in the extent or timing of gene expression can have profound effects on molecular and organismal phenotypes and thereby drive evolution (King and Wilson 1975;Wray 2007). Heritable noncoding variation can alter gene expression in cis or in trans (Skelly et al. 2009), and both have been shown to contribute significantly to gene expression variation (Ronald et al. 2005;Tirosh et al. 2009;Skelly et al. 2011). Two primary mechanisms by which cis variation can increase gene expression are increases in gene copy number and point mutations in regulatory regions. However, the relative effect size of amplification compared to point mutation is not known.It has been proposed that amplification is both quickly achieved and then reverts after fixation of fitness-increasing point mutations (Hendrickson et al. 2002;Yona et al. 2012). This mechanism raises the question of whether amplification is simply a transitional state that provides an increased chance for a beneficial point mutation to occur. Alternatively, gene amplification may be so frequently observed because the fitness effects it confers are greater than those achievable by point mutations. Gene amplifications have been found to be advantageous in many contexts, including phenotypic evolution (Hoekstra and Coyne 2007;Stern and Orgogozo 2008) an...

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Section: Discussionmentioning

confidence: 99%

Comprehensive Analysis of the SUL1 Promoter of Saccharomyces cerevisiae

et al. 2016

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“…Additional double-gene deletions were generated using a previously described protocol [30]. SK1 wild type and deletion strains were obtained from Wilkening et al [31]. Strains were phenotyped in YPD, YPD+Menadione (50μM) and YPD+CdCl 2 (10μM).…”

Section: Methodsmentioning

confidence: 99%

The Modular Adaptive Ribosome

et al. 2016

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The ribosome is an ancient machine, performing the same function across organisms. Although functionally unitary, recent experiments suggest specialized roles for some ribosomal proteins. Our central thesis is that ribosomal proteins function in a modular fashion to decode genetic information in a context dependent manner. We show through large data analyses that although many ribosomal proteins are essential with consistent effect on growth in different conditions in yeast and similar expression across cell and tissue types in mice and humans, some ribosomal proteins are used in an environment specific manner. The latter set of variable ribosomal proteins further function in a coordinated manner forming modules, which are adapted to different environmental cues in different organisms. We show that these environment specific modules of ribosomal proteins in yeast have differential genetic interactions with other pathways and their 5’UTRs show differential signatures of selection in yeast strains, presumably to facilitate adaptation. Similarly, we show that in higher metazoans such as mice and humans, different modules of ribosomal proteins are expressed in different cell types and tissues. A clear example is nervous tissue that uses a ribosomal protein module distinct from the rest of the tissues in both mice and humans. Our results suggest a novel stratification of ribosomal proteins that could have played a role in adaptation, presumably to optimize translation for adaptation to diverse ecological niches and tissue microenvironments.

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“…These approaches have recently been applied to yeast to map quantitative trait loci (QTL) responsible for industrially important traits such as sulfite resistance [90], aromatic compound production [91], flocculation [92], and thermotolerance [93]. Future work on other domesticated microbes would first have to focus on establishing conditions to induce sexual reproduction in cryptically sexual food-related eukaryotic microbes [94], as has been initiated in Aspergillus oryzae [95,96] and Penicillium roqueforti [97,98].…”

Section: Future Directionsmentioning

confidence: 99%

The genomics of microbial domestication in the fermented food environment

Gibbons

Rinker

2015

Current Opinion in Genetics & Development

117

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Shortly after the agricultural revolution, the domestication of bacteria, yeasts, and molds, played an essential role in enhancing the stability, quality, flavor, and texture of food products. These domestication events were likely the result of human food production practices that entailed the continual recycling of isolated microbial communities in the presence of abundant agricultural food sources. We suggest that within these novel agrarian food niches the metabolic requirements of those microbes became regular and predictable resulting in rapid genomic specialization through such mechanisms as pseudogenization, genome decay, interspecific hybridization, gene duplication, and horizontal gene transfer. The ultimate result was domesticated strains of microorganisms with enhanced fermentative capacities.

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An Evaluation of High-Throughput Approaches to QTL Mapping in Saccharomyces cerevisiae

Cited by 79 publications

References 77 publications

Comprehensive Analysis of the SUL1 Promoter of Saccharomyces cerevisiae

Comprehensive Analysis of the SUL1 Promoter of Saccharomyces cerevisiae

The Modular Adaptive Ribosome

The genomics of microbial domestication in the fermented food environment

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