Loyola Marymount University Acclimation of Saccharomyces cerevisiae to Low Temperature: A Chemostat-based Transcriptome Analysis Tai, S. L., Daran-Lapujade, P., Walsh, M. C., Pronk, J. T., & Daran, J. M. (2007) Molecular Biology of the Cell, 18, 5100-5112. Student Names Department of Biology Loyola Marymount University BIOL 478 April 22, 2014

Outline Importance of chemostat cultures Separation of variables Adaptation vs. acclimation Viability of S. cerevisiae in experimental conditions Environment dependent temperature-responsive genes Identification Function Accumulation and regulation of storage carbohydrates Promoter analysis Adaptation and acclimation Growth rate and gene expression Environmental stress response genes

Outline Importance of chemostat cultures Separation of variables Adaptation vs. acclimation Viability of S. cerevisiae in experimental conditions Environment dependent temperature-responsive genes Identification Function Accumulation and regulation of storage carbohydrates Promoter analysis Adaptation and acclimation Growth rate and gene expression Environmental stress response genes

Chemostat cultures allow for fixed experimental conditions Chemostat culture: chemical environment is static as medium is constantly being added and removed Allows for fine control of variables Specific growth rate Some genes are context dependent Adaptation: a rapid, highly dynamic stress-response phenomena Acclimation: establishment of a physiological state in which regulatory mechanisms have resulted in full adaptation of genome expression to the environmental conditions

Outline Importance of chemostat cultures Separation of variables Adaptation vs. acclimation Viability of S. cerevisiae in experimental conditions Environment dependent temperature-responsive genes Identification Function Accumulation and regulation of storage carbohydrates Promoter analysis Adaptation and acclimation Growth rate and gene expression Environmental stress response genes

Viability of S. Cerevisiae grown in four conditions Showing physiological traits of the yeast in all 4 different conditions Yeast is viable at all temperatures in all conditions Similar values show that growth not strictly affected by growth temperature

Outline Importance of chemostat cultures Separation of variables Adaptation vs. acclimation Viability of S. cerevisiae in experimental conditions Environment dependent temperature-responsive genes Identification Function Accumulation and regulation of storage carbohydrates Promoter analysis Adaptation and acclimation Growth rate and gene expression Environmental stress response genes

Environment Dependent Temperature-responsive Genes Over 1000 genes were found to have a change in transcription level Genes not consistently transcribed across the different conditions

Screening of Temperature-responsive Genes for Function Wanted to find regulatory networks that linked genes Grouped together to see if there were any similarities within their functional groups Observed changes in Transporters of limiting nutrient Translational machinery NCR genes and catabolite repression

Outline Importance of chemostat cultures Separation of variables Adaptation vs. acclimation Viability of S. cerevisiae in experimental conditions Environment dependent temperature-responsive genes Identification Function Accumulation and regulation of storage carbohydrates Promoter analysis Adaptation and acclimation Growth rate and gene expression Environmental stress response genes

Storage Carbohydrate Accumulation and Regulation Trehalose and glycogen as markers of cold shock Different levels in different conditions (12 vs. 30 C˚) Accumulation and transcription of trehalose and glycogen not needed in steady state cold stress More proteins at 12 C˚ than 30 C˚

Outline Importance of chemostat cultures Separation of variables Adaptation vs. acclimation Viability of S. cerevisiae in experimental conditions Environment dependent temperature-responsive genes Identification Function Accumulation and regulation of storage carbohydrates Promoter analysis Adaptation and acclimation Growth rate and gene expression Environmental stress response genes

Promoter Analysis Reveals Trends in Gene Regulation STRE elements in promoters of downregulated genes in N-limited cultures Upregulation in both conditions show PAC regulatory motifs in promoters Common promoter sequences show possible regulation relationships

Outline Importance of chemostat cultures Separation of variables Adaptation vs. acclimation Viability of S. cerevisiae in experimental conditions Environment dependent temperature-responsive genes Identification Function Accumulation and regulation of storage carbohydrates Promoter analysis Adaptation and acclimation Growth rate and gene expression Environmental stress response genes

Comparison of Data Sets—Adaptation Sahara et al. 2002, Schade et al. 2004, and Murata et al. 2006 Found 259 genes that all responded, but not consistent Context dependent regulation

Adaptation and Acclimation Chemostat study compared with Sahara et al. 2002, Schade et al. 2004, and Murata et al. 2006 29 genes total were in common, only 11 showed consistency

Outline Importance of chemostat cultures Separation of variables Adaptation vs. acclimation Viability of S. cerevisiae in experimental conditions Environment dependent temperature-responsive genes Identification Function Accumulation and regulation of storage carbohydrates Promoter analysis Adaptation and acclimation Growth rate and gene expression Environmental stress response genes

Effect of Growth Rate on Gene Expression Genes consistently regulated in batch studies compared to genes that transcriptionally responded to growth rates (Castrillo and Regenberg) Genes consistently regulated in chemostat study compared to Castrillo and Regenberg Growth rate as affecting factor

Outline Importance of chemostat cultures Separation of variables Adaptation vs. acclimation Viability of S. cerevisiae in experimental conditions Environment dependent temperature-responsive genes Identification Function Accumulation and regulation of storage carbohydrates Promoter analysis Adaptation and acclimation Growth rate and gene expression Environmental stress response genes

Environmental Stress Response Gene Comparison ESR: environmental stress response General mechanism responds to multiple stimuli Batch and chemostat genes compared with Gasch et al. 2000 Some genes showed opposite results

Summary Chemostat cultures allow for separation of variables and can highlight condition-dependent transcription S. cerevisiae able to grow under multiple conditions with different gene expression Environment dependent temperature-responsive genes have similar functional groups Accumulation and regulation of storage carbohydrates not present in long term exposure to cold stress Promoter analysis reveals that functional groups of genes have similar promoters Adaptation and acclimation have different gene sets associated with them Growth rate can affect how genes are expressed Environmental stress response genes different in batch vs. chemostat studies

Acknowledgments Sammi Flores