Project description:Resistance to agricultural fungicides in the field has created a need for discovering fungicides with new modes of action. DNA microarrays, because they provide information on expression of many genes simultaneously, could help to identify the modes of action. To begin an expression pattern database for agricultural fungicides, transcriptional patterns of Saccharomyces cerevisiae strain S288C genes were analysed following 2-h treatments with I50 concentrations of ergosterol biosynthesis inhibitors commonly used against plant pathogenic fungi. Eight fungicides, representing three classes of ergosterol biosynthesis inhibitors, were tested. To compare gene expression in response to a fungicide with a completely different mode of action, a putative methionine biosynthesis inhibitor (MBI) was also tested. Expression patterns of ergosterol biosynthetic genes supported the roles of Class I and Class II inhibitors in affecting ergosterol biosynthesis, confirmed that the putative MBI did not affect ergosterol biosynthesis, and strongly suggested that in yeast, the Class III inhibitor did not affect ergosterol biosynthesis. The MBI affected transcription of three genes involved in methionine metabolism, whereas there were essentially no effects of ergosterol synthesis inhibitors on methionine metabolism genes. There were no consistent patterns in other up- or downregulated genes between fungicides. These results suggest that inspection of gene response patterns within a given pathway may serve as a useful first step in identifying possible modes of action of fungicides. agricultural sterol biosynthesis inhibitor fungicides. Keywords = agriculture Keywords = ergosterol Keywords = methionine Keywords = fungicide Keywords = Saccharomyces cerevisiae S288C Keywords = biosynthesis
Project description:Resistance to agricultural fungicides in the field has created a need for discovering fungicides with new modes of action. DNA microarrays, because they provide information on expression of many genes simultaneously, could help to identify the modes of action. To begin an expression pattern database for agricultural fungicides, transcriptional patterns of Saccharomyces cerevisiae strain S288C genes were analysed following 2-h treatments with I50 concentrations of ergosterol biosynthesis inhibitors commonly used against plant pathogenic fungi. Eight fungicides, representing three classes of ergosterol biosynthesis inhibitors, were tested. To compare gene expression in response to a fungicide with a completely different mode of action, a putative methionine biosynthesis inhibitor (MBI) was also tested. Expression patterns of ergosterol biosynthetic genes supported the roles of Class I and Class II inhibitors in affecting ergosterol biosynthesis, confirmed that the putative MBI did not affect ergosterol biosynthesis, and strongly suggested that in yeast, the Class III inhibitor did not affect ergosterol biosynthesis. The MBI affected transcription of three genes involved in methionine metabolism, whereas there were essentially no effects of ergosterol synthesis inhibitors on methionine metabolism genes. There were no consistent patterns in other up- or downregulated genes between fungicides. These results suggest that inspection of gene response patterns within a given pathway may serve as a useful first step in identifying possible modes of action of fungicides. agricultural sterol biosynthesis inhibitor fungicides. Keywords = agriculture Keywords = ergosterol Keywords = methionine Keywords = fungicide Keywords = Saccharomyces cerevisiae S288C Keywords = biosynthesis
Project description:Correlations between the chemical structures of agricultural fungicides and mRNA expression levels following exposure of Saccharomyces cerevisiae to toxic doses of thiuram, zineb, maneb, TPN, and PCP were examined. Structurally, thiuram, zineb, and maneb are dithiocarbamate fungicides, whereas TPN and PCP are not. To characterize chemical toxicity, genes expression was classified according to the functional groups used by the MIPS database. However, no correlations between the classification scheme and chemical structures were found. Hierarchical clustering of gene expression profiles was performed to characterize the effects of the five chemicals. According to this analysis the similarity of gene expression profiles depended on the similarity of chemical structures. These results suggest that DNA microarray technology has potential for predicting the major chemicals which will cause environmental toxicity and will provide information on new biomonitoring methods. Keywords: stress response
Project description:Ergosterol is a prominent component of the yeast plasma membrane and essential for yeast cell viability. It is synthesized in the endoplasmic reticulum and transported to the plasma membrane by non-vesicular mechanisms requiring carrier proteins. Oxysterol binding protein homologues and yeast StARkin proteins have been proposed to function as sterol carriers. Although many of these proteins are capable of transporting sterols between synthetic lipid vesicles in vitro, they are not essential for ergosterol transport in cells, indicating that they may be functionally redundant with each other or with additional - as yet unidentified - sterol carriers. To address this point we hypothesized that sterol transport proteins are also sterol binding proteins (SBPs), and used an in vitro chemo-proteomic strategy to identify all cytosolic SBPs. We generated a cytosol fraction enriched in SBPs and captured the proteins with a photoreactive clickable cholesterol analog. Quantitative proteomics of the captured proteins identified 342 putative SBPs. Analysis of these identified proteins based on their annotated function, reported drug phenotypes, interactions with proteins regulating lipid metabolism, gene ontology, and presence of mammalian orthologs revealed a subset of 62 characterized and nine uncharacterized candidates. Five of the uncharacterized proteins play a role in maintaining plasma membrane integrity as their absence affects the ability of cells to grow in the presence of nystatin or myriocin. We anticipate that the dataset reported here will be a comprehensive resource for functional analysis of sterol binding/transport proteins and provide insights into novel aspects of non-vesicular sterol trafficking.
Project description:During fermentation Saccharomyces yeast produces various aroma-active metabolites determining the different characteristics of aroma and taste in fermented beverages. Amino acid utilization by yeast during brewer´s wort fermentation is seen as linked to flavour profile. To better understand the relationship between the biosynthesis of aroma relevant metabolites and the importance of amino acids, DNA microarrays were performed for Saccharomyces cerevisiae strain S81 and Saccharomyces pastorianus var. carlsbergensis strain S23, respectively. Thereby, changes in transcription of genes were measured, which are associated with amino acid assimilation and its derived aroma-active compounds during fermentation.