A Systems Approach Demonstrating Sphingolipid-Dependent Transcription in Stress Responses

Microarray hybridization allows genome-wide screening of changes in mRNA levels under stress conditions. In Saccharomyces cerevisiae , this approach has demonstrated that responses to heat stress, oxidative stress, nutrient deprivation, and other stress signals are highly overlapping and mRNA levels of a core group of genes, termed ‘Environmental Stress Response’ (ESR) genes, respond similarly to many stressors. In addition to changes in mRNA levels, stress responses induce wide changes in cell metabolic pathways and metabolite levels. Microarrays coupled with chemical inhibition of these pathways and/or using organisms with genetic mutations in enzymes in the pathways of interest allow determination of the roles of specific metabolites in gene expression. In cases where high-throughput ‘-omics’ strategies are available for determining changes in a spectrum of metabolites, these datasets can be integrated with gene expression data to obtain a systems view of regulations and functions of a given pathway. We have used these approaches to determine the regulation and functions of sphingolipid synthesis in Saccharomyces cerevisiae . Microarray hybridization and sphingolipidomic analysis experiments were performed on two yeast strains bearing mutations in enzymes of sphingolipid metabolism (and their respective parental strains), under normal conditions and during heat stress. These strategies have revealed diverse roles for sphingolipids in regulating stress response genes, and moreover, could be applied to numerous biological systems and thus provide a method to elucidate activities for a vast array of biomolecules, the metabolic pathways by which they are generated, and their cellular functions.