Project description:The Saccharomyces cerevisiae gene PIF1 encodes a conserved eukaryotic DNA helicase required for both mitochondrial and nuclear DNA integrity. Our previous work revealed that a pif1D strain is tolerant to zinc overload. We demonstrate here that this effect is independent of the Pif1 helicase activity and is only observed when the protein is absent from the mitochondria. pif1 cells accumulate abnormal amounts of mitochondrial zinc and iron. Transcriptional profiling reveals that pif1 cells under standard growth conditions overexpress aconitase-related genes. When exposed to zinc, pif1 cells show lower induction of genes encoding iron (siderophores) transporters and higher expression of genes related to oxidative stress responses than wild type cells. Coincidently, pif1 mutants are less prone to zinc-induced oxidative stress and display a higher reduced/oxidized glutathione ratio. Strikingly, although pif1 cells contain normal amounts of the Aco1 protein, they completely lack aconitase activity. Loss of Aco1 activity is also observed when the cell expresses a non-mitochondrially targeted form of Pif1. We postulate that lack of Pif1 forces aconitase to play its DNA protective role as nucleoid protein and that this would trigger a domino effect on iron homeostasis resulting in increased zinc tolerance.
Project description:The Saccharomyces cerevisiae gene PIF1 encodes a conserved eukaryotic DNA helicase required for both mitochondrial and nuclear DNA integrity. Our previous work revealed that a pif1D strain is tolerant to zinc overload. We demonstrate here that this effect is independent of the Pif1 helicase activity and is only observed when the protein is absent from the mitochondria. pif1 cells accumulate abnormal amounts of mitochondrial zinc and iron. Transcriptional profiling reveals that pif1 cells under standard growth conditions overexpress aconitase-related genes. When exposed to zinc, pif1 cells show lower induction of genes encoding iron (siderophores) transporters and higher expression of genes related to oxidative stress responses than wild type cells. Coincidently, pif1 mutants are less prone to zinc-induced oxidative stress and display a higher reduced/oxidized glutathione ratio. Strikingly, although pif1 cells contain normal amounts of the Aco1 protein, they completely lack aconitase activity. Loss of Aco1 activity is also observed when the cell expresses a non-mitochondrially targeted form of Pif1. We postulate that lack of Pif1 forces aconitase to play its DNA protective role as nucleoid protein and that this would trigger a domino effect on iron homeostasis resulting in increased zinc tolerance. Four samples were analyzed: WT and pif1 mutant both, in the presence and in the absence of 5 mM ZnCl2 for 1 h. We compared the expression profile of: 1) WT+Zn vs WT-Zn, 2) pif1 mutant+Zn vs pif1 mutant-Zn, 3) pif1 mutant vs WT, and 4) pif1 mutant+Zn vs WT+Zn. A Dye-swap was carried out for each comparison of samples. Total number of chips analyzed: 8.
Project description:Industrial bioethanol production may involve a low pH environment,improving the tolerance of S. cerevisiae to a low pH environment caused by inorganic acids may be of industrial importance to control bacterial contamination, increase ethanol yield and reduce production cost. Through analysis the transcriptomic data of Saccharomyces cerevisiae with different ploidy under low pH stress, we hope to find the tolerance mechanism of Saccharomyces cerevisiae to low pH.
Project description:High concenHigh concentration acetic acid in the fermentation medium represses cell growth, metabolism and fermentation efficiency of Saccharomyces cerevisiae, which is widely used for cellulosic ethanol production. Our previous study proved that supplementation of zinc sulfate in the fermentation medium improved cell growth and ethanol fermentation performance of S. cerevisiae under acetic acid stress condition. However, the molecular mechanisms is still unclear. To explore the underlying mechanism of zinc sulfate protection against acetic acid stress, transcriptomic and proteomic analysis were performed. The changed genes and proteins are related to carbon metabolism, amino acid biosynthesis, energy metabolism, vitamin biosynthesis and stress responses. In a total, 28 genes showed same expression in transcriptomic and proteomic data, indicating that zinc sulfate affects gene expression at posttranscriptional and posttranslational levels.tration acetic acid in the fermentation medium represses cell growth, metabolism and fermentation efficiency of Saccharomyces cerevisiae, which is widely used for cellulosic ethanol production. Our previous study proved that supplementation of zinc sulfate in the fermentation medium improved cell growth and ethanol fermentation performance of S. cerevisiae under acetic acid stress condition. However, the molecular mechanisms is still unclear. To explore the underlying mechanism of zinc sulfate protection against acetic acid stress, transcriptomic and proteomic analysis were performed. The changed genes and proteins are related to carbon metabolism, amino acid biosynthesis, energy metabolism, vitamin biosynthesis and stress responses. In a total, 28 genes showed same expression in transcriptomic and proteomic data, indicating that zinc sulfate affects gene expression at posttranscriptional and posttranslational levels.
Project description:Transcriptional profiling of Saccharomyces cerevisiae cells comparing the W303-1A wildtype with the W303-1A double mutant for MSN2 and MSN4 during zinc deficient conditions Keywords: Genetic modification with zinc limitation
Project description:The vanillin tolerance Saccharomyces cerevisiae was screened and compared intracellular ergosterol levels with several laboratory yeast strains, to study potential relationship between ergosterol contents and vanillin tolerance. S. cerevisiae NBRC1950 was selected as a vanillin tolerant strain. Its ergosterol contents were higher than those of laboratory strains. The results of DNA microarray and quantitative RT-PCR analysis showed that 5 genes involved in ergosterol biosynthesis (ERG28, HMG1, MCR1, ERG5 and ERG7) were up-regulated in NBRC 1950 compared with strain X2180, suggested that high expressions of genes involved in ergosterol biosynthesis may cause for the high ergosterol content in strain NBRC 1950. S. cerevisiae HX strain, which was a high ergosterol content strain derived from X2180, became more tolerant to vanillin compared with the parental strain. It is suggested that high ergosterol contents may be in part responsible for vanillin tolerance. These findings provide a biotechnological basis for the molecular engineering of S. cerevisiae with increased tolerance to vanillin.
Project description:Metabolic engineering of Saccharomyces cerevisiae for efficient monoterpenes production was mostly restricted by the limited tolerance to these chemicals. Understanding of the molecular mechanisms underlying the tolerance of S. cerevisiae to monoterpenes was essential for the de novo biosynthesis these chemicals in S. cerevisiae. In this study, commercial oligonucleotide microarray assays were performed to investigate the global response of S. cerevisiae to typical monoterpene D-limonene under transcriptional level. Yeast cell treated with sublethal dose of D-liomonene, gene change profiles were investigated at transcription level and the microarry data were also verified with quantitative real time PCR.
Project description:Transcriptional profiling of Saccharomyces cerevisiae cells comparing the W303-1A wildtype with the W303-1A double mutant for MSN2 and MSN4 during zinc deficient conditions Keywords: Genetic modification with zinc limitation Two condition experiment, W303-1A vs W303-1A delta MSN2, MSN4. Biological replicates: 2 wildtype, 2 knock-out, independently grown and harvested.