Audrey P. Gasch

The genomics of yeast responses to environmental stress and starvation

Abstract. Unicellular organisms such as yeast have evolved to survive constant fluctuations in their external surroundings by rapidly adapting their internal systems to meet the challenges of each new environment. One aspect of this cellular adaptation is the reorganization of genomic expression to the program required for growth in each environment. The reprogramming of genomic expression can be unveiled using DNA microarrays, which measure the relative transcript abundance of essentially every gene in an organisms genome. Characterizing environmentally triggered gene expression changes provides insights into when, where, and how each gene is expressed and offers a glimpse at the physiological response of the cells to changes in their surroundings. This review will focus on the genomic expression responses of the budding yeast Saccharomyces cerevisiae to diverse environmental changes, highlighting some of the themes that have emerged from the collection of published yeast genomic expression studies. The results of these studies present insights as to how yeast cells sense and respond to each new environment, and suggest mechanisms that this organism uses to survive stressful environmental changes.

Conservation and evolution of cis-regulatory systems in ascomycete fungi

Relatively little is known about the mechanisms through which gene expression regulation evolves. To investigate this, we systematically explored the conservation of regulatory networks in fungi by examining the cis-regulatory elements that govern the expression of coregulated genes. We first identified groups of coregulated Saccharomyces cerevisiae genes enriched for genes with known upstream or downstream cis-regulatory sequences. Reasoning that many of these gene groups are coregulated in related species as well, we performed similar analyses on orthologs of coregulated S. cerevisiae genes in 13 other ascomycete species. We find that many species-specific gene groups are enriched for the same flanking regulatory sequences as those found in the orthologous gene groups from S. cerevisiae, indicating that those regulatory systems have been conserved in multiple ascomycete species. In addition to these clear cases of regulatory conservation, we find examples of cis-element evolution that suggest multiple modes of regulatory diversification, including alterations in transcription factor-binding specificity, incorporation of new gene targets into an existing regulatory system, and cooption of regulatory systems to control a different set of genes. We investigated one example in greater detail by measuring the in vitro activity of the S. cerevisiae transcription factor Rpn4p and its orthologs from Candida albicans and Neurospora crassa. Our results suggest that the DNA binding specificity of these proteins has coevolved with the sequences found upstream of the Rpn4p target genes and suggest that Rpn4p has a different function in N. crassa.

A dynamic model of proteome changes reveals new roles for transcript alteration in yeast.

The transcriptome and proteome change dynamically as cells respond to environmental stress; however, prior proteomic studies reported poor correlation between mRNA and protein, rendering their relationships unclear. To address this, we combined high mass accuracy mass spectrometry with isobaric tagging to quantify dynamic changes in ∼2500 Saccharomyces cerevisiae proteins, in biological triplicate and with paired mRNA samples, as cells acclimated to high osmolarity. Surprisingly, while transcript induction correlated extremely well with protein increase, transcript reduction produced little to no change in the corresponding proteins. We constructed a mathematical model of dynamic protein changes and propose that the lack of protein reduction is explained by cell‐division arrest, while transcript reduction supports redistribution of translational machinery. Furthermore, the transient ‘burst’ of mRNA induction after stress serves to accelerate change in the corresponding protein levels. We identified several classes of post‐transcriptional regulation, but show that most of the variance in protein changes is explained by mRNA. Our results present a picture of the coordinated physiological responses at the levels of mRNA, protein, protein‐synthetic capacity, and cellular growth.

Variations in stress sensitivity and genomic expression in diverse S. cerevisiae isolates.

Interactions between an organism and its environment can significantly influence phenotypic evolution. A first step toward understanding this process is to characterize phenotypic diversity within and between populations. We explored the phenotypic variation in stress sensitivity and genomic expression in a large panel of Saccharomyces strains collected from diverse environments. We measured the sensitivity of 52 strains to 14 environmental conditions, compared genomic expression in 18 strains, and identified gene copy-number variations in six of these isolates. Our results demonstrate a large degree of phenotypic variation in stress sensitivity and gene expression. Analysis of these datasets reveals relationships between strains from similar niches, suggests common and unique features of yeast habitats, and implicates genes whose variable expression is linked to stress resistance. Using a simple metric to suggest cases of selection, we found that strains collected from oak exudates are phenotypically more similar than expected based on their genetic diversity, while sake and vineyard isolates display more diverse phenotypes than expected under a neutral model. We also show that the laboratory strain S288c is phenotypically distinct from all of the other strains studied here, in terms of stress sensitivity, gene expression, Ty copy number, mitochondrial content, and gene-dosage control. These results highlight the value of understanding the genetic basis of phenotypic variation and raise caution about using laboratory strains for comparative genomics.

Comparative genomics of the environmental stress response in ascomycete fungi

Unicellular fungi thrive in diverse niches around the world, and many of these niches present unique and stressful challenges that must be contended with by their inhabitants. Numerous studies have investigated the genomic expression responses to environmental stress in ‘model’ ascomycete fungi, including Saccharomyces cerevisiae, Candida albicans and Schizosaccharomyces pombe. This review presents a comparative‐genomics perspective on the environmental stress response, a common response to diverse stresses. Implications for the role of this response, based on its presence or absence in fungi from disparate ecological niches, are discussed. Copyright

Comparative genomics of xylose-fermenting fungi for enhanced biofuel production

Cellulosic biomass is an abundant and underused substrate for biofuel production. The inability of many microbes to metabolize the pentose sugars abundant within hemicellulose creates specific challenges for microbial biofuel production from cellulosic material. Although engineered strains of Saccharomyces cerevisiae can use the pentose xylose, the fermentative capacity pales in comparison with glucose, limiting the economic feasibility of industrial fermentations. To better understand xylose utilization for subsequent microbial engineering, we sequenced the genomes of two xylose-fermenting, beetle-associated fungi, Spathaspora passalidarum and Candida tenuis. To identify genes involved in xylose metabolism, we applied a comparative genomic approach across 14 Ascomycete genomes, mapping phenotypes and genotypes onto the fungal phylogeny, and measured genomic expression across five Hemiascomycete species with different xylose-consumption phenotypes. This approach implicated many genes and processes involved in xylose assimilation. Several of these genes significantly improved xylose utilization when engineered into S. cerevisiae, demonstrating the power of comparative methods in rapidly identifying genes for biomass conversion while reflecting on fungal ecology.

The environmental stress response: a common yeast response to diverse environmental stresses

Unicellular organisms require specific internal conditions for optimal growth and function, however sudden changes in the external environment can perturb the internal milieu, disrupting normal processes. Therefore, cells must maintain their internal system despite fluctuations in the external surroundings. One mechanism that yeast cells use to protect the internal system from the effects of environmental variation is to initiate a common gene expression program that generally protects the cell during stressful times. This program, referred to as the environmental stress response, includes ∼900 genes whose expression is stereotypically altered when yeast cells are shifted to stressful environments. The coordinated expression changes of these genes is a common feature of the responses to many different environments, however the regulation of these expression changes is gene-specific and condition-specific, indicating that initiation of the program is precisely controlled in response to each new environment. This review will focus on recent developments in defining and characterizing the genes that participate in the environmental stress response and the regulatory mechanisms that the cell utilizes to orchestrate this program.

The histone deacetylase Rpd3p is required for transient changes in genomic expression in response to stress

BackgroundYeast responding to stress activate a large gene expression program called the Environmental Stress Response that consists of approximately 600 repressed genes and approximately 300 induced genes. Numerous factors are implicated in regulating subsets of Environmental Stress Response genes; however, a complete picture of Environmental Stress Response regulation remains unclear. We investigated the role of the histone deacetylase Rpd3p, previously linked to the upstream regions of many Environmental Stress Response genes, in producing Environmental Stress Response gene expression changes in response to stress.ResultsWe found that the Rpd3-Large complex is required for proper expression of both induced and repressed Environmental Stress Response genes under multiple stress conditions. Cells lacking RPD3 or the Rpd3-Large subunit PHO23 had a major defect in Environmental Stress Response initiation, particularly during the transient phase of expression immediately after stress exposure. Chromatin-immunoprecipitation showed a direct role for Rpd3-Large at representative genes; however, there were different effects on nucleosome occupancy and histone deacetylation at different promoters. Computational analysis implicated regulators that may act with Rpd3p at Environmental Stress Response genes. We provide genetic and biochemical evidence that Rpd3p is required for binding and action of the stress-activated transcription factor Msn2p, although the contribution of these factors differs for different genes.ConclusionsOur results implicate Rpd3p as an important co-factor in the Environmental Stress Response regulatory network, and suggest the importance of histone modification in producing transient changes in gene expression triggered by stress.

Exploiting the yeast stress-activated signaling network to inform on stress biology and disease signaling

Healthy cells utilize intricate systems to monitor their environment and mount robust responses in the event of cellular stress. Whether stress arises from external insults or defects due to mutation and disease, cells must be able to respond precisely to mount the appropriate defenses. Multi-faceted stress responses are generally coupled with arrest of growth and cell-cycle progression, which both limits the transmission of damaged materials and serves to reallocate limited cellular resources toward defense. Therefore, stress defense versus rapid growth represent competing interests in the cell. How eukaryotic cells set the balance between defense versus proliferation, and in particular knowledge of the regulatory networks that control this decision, are poorly understood. In this perspective, we expand upon our recent work inferring the stress-activated signaling network in budding yeast, which captures pathways controlling stress defense and regulators of growth and cell-cycle progression. We highlight similarities between the yeast and mammalian stress responses and explore how stress-activated signaling networks in yeast can inform on signaling defects in human cancers.

Multiple Means to the Same End: The Genetic Basis of Acquired Stress Resistance in Yeast

In nature, stressful environments often occur in combination or close succession, and thus the ability to prepare for impending stress likely provides a significant fitness advantage. Organisms exposed to a mild dose of stress can become tolerant to what would otherwise be a lethal dose of subsequent stress; however, the mechanism of this acquired stress tolerance is poorly understood. To explore this, we exposed the yeast gene-deletion libraries, which interrogate all essential and non-essential genes, to successive stress treatments and identified genes necessary for acquiring subsequent stress resistance. Cells were exposed to one of three different mild stress pretreatments (salt, DTT, or heat shock) and then challenged with a severe dose of hydrogen peroxide (H2O2). Surprisingly, there was little overlap in the genes required for acquisition of H2O2 tolerance after different mild-stress pretreatments, revealing distinct mechanisms of surviving H2O2 in each case. Integrative network analysis of these results with respect to protein–protein interactions, synthetic–genetic interactions, and functional annotations identified many processes not previously linked to H2O2 tolerance. We tested and present several models that explain the lack of overlap in genes required for H2O2 tolerance after each of the three pretreatments. Together, this work shows that acquired tolerance to the same severe stress occurs by different mechanisms depending on prior cellular experiences, underscoring the context-dependent nature of stress tolerance.