ABSTRACT: Systematic identification of regulators of oxidative stress reveals non-canonical roles for peroxisomal import and the pentose phosphate pathway
Project description:Systematic identification of regulators of oxidative stress reveals non-canonical roles for peroxisomal import and the pentose phosphate pathway
Project description:In industrial fermentations of Saccharomyces cerevisiae, transient changes in oxygen concentration commonly occur and it is important to understand the behaviour of cells during these changes. Saccharomyces cerevisiae CEN.PK113-1A was grown in glucose-limited chemostat culture with 1.0% and 20.9% O2 in the inlet gas (D= 0.10 /h, pH5, 30C). After steady state was achieved, oxygen was replaced with nitrogen and cultures were followed until new steady state was achieved. The overall responses to anaerobic conditions of cells initially in different conditions were very similar. Independent of initial culture conditions, transient downregulation of genes related to growth and cell proliferation, mitochondrial translation and protein import, and sulphate assimilation was seen. In addition, transient or permanent upregulation of genes related to protein degradation, and phosphate and amino acid uptake was observed in all cultures. However, only in the initially oxygen-limited cultures was a transient upregulation of genes related to fatty acid oxidation, peroxisomal biogenesis, oxidative phosphorylation, TCA cycle, response to oxidative stress, and pentose phosphate pathway observed. Furthermore, from the initially oxygen-limited conditions, a rapid response around the metabolites of upper glycolysis and the pentose phosphate pathway was seen, while from the initially fully aerobic conditions, a slower response around the pathways for utilisation of respiratory carbon sources was observed. Time series analysis starting from two (1% and 20.9%) levels of oxygen provision. Seven timepoints from both time series and two biological replicates from each timepoint were analysed.
Project description:In industrial fermentations of Saccharomyces cerevisiae, transient changes in oxygen concentration commonly occur and it is important to understand the behaviour of cells during these changes. Saccharomyces cerevisiae CEN.PK113-1A was grown in glucose-limited chemostat culture with 1.0% and 20.9% O2 in the inlet gas (D= 0.10 /h, pH5, 30C). After steady state was achieved, oxygen was replaced with nitrogen and cultures were followed until new steady state was achieved. The overall responses to anaerobic conditions of cells initially in different conditions were very similar. Independent of initial culture conditions, transient downregulation of genes related to growth and cell proliferation, mitochondrial translation and protein import, and sulphate assimilation was seen. In addition, transient or permanent upregulation of genes related to protein degradation, and phosphate and amino acid uptake was observed in all cultures. However, only in the initially oxygen-limited cultures was a transient upregulation of genes related to fatty acid oxidation, peroxisomal biogenesis, oxidative phosphorylation, TCA cycle, response to oxidative stress, and pentose phosphate pathway observed. Furthermore, from the initially oxygen-limited conditions, a rapid response around the metabolites of upper glycolysis and the pentose phosphate pathway was seen, while from the initially fully aerobic conditions, a slower response around the pathways for utilisation of respiratory carbon sources was observed.
Project description:Exposure to oxygen and light generates photooxidative stress by the bacteriochlorophyll a mediated formation of singlet oxygen (1O2) in the facultative photosynthetic bacterium Rhodobacter sphaeroides. We have identified SorY as an sRNA, which is induced under several stress conditions and confers increased resistance against 1O2. SorY by direct interaction decreases the levels of takP mRNA, encoding a TRAP-T transporter. A takP mutant shows higher resistance to 1O2 than the wild type, which is no longer affected by SorY. We present a model in which SorY reduces the metabolite flux into the TCA cycle by reducing malate import through TakP. It was previously shown that oxidative stress in bacteria leads to switch from glycolysis to the pentose phosphate cycle and to reduced activity of the tricaboxylic acid cycle. As a consequence the production of the prooxidant NADH is reduced and production of the protective NADPH is enhanced. In R. sphaeroides enzymes for glycolysis, pentose phosphate pathway, EntnerM-bM-^@M-^SDoudoroff pathway and gluconeogenesis are induced in response to 1O2 by the alternative sigma factor RpoHII. The same is true for the sRNA SorY. By limiting malate import SorY thus contributes to the balance of the metabolic fluxes under photooxidative stress conditions. This assigns a so far unknown function to an sRNA in oxidative stress response. RNA samples collected from a control strain harbouring an empty vector (2.4.1pBBR) and of the SorY overexpressing strain (2.4.1pBBRSorYi) after 10 min of 1O2 stress were analyzed by two-color microarrays
Project description:Exposure to oxygen and light generates photooxidative stress by the bacteriochlorophyll a mediated formation of singlet oxygen (1O2) in the facultative photosynthetic bacterium Rhodobacter sphaeroides. We have identified SorY as an sRNA, which is induced under several stress conditions and confers increased resistance against 1O2. SorY by direct interaction decreases the levels of takP mRNA, encoding a TRAP-T transporter. A takP mutant shows higher resistance to 1O2 than the wild type, which is no longer affected by SorY. We present a model in which SorY reduces the metabolite flux into the TCA cycle by reducing malate import through TakP. It was previously shown that oxidative stress in bacteria leads to switch from glycolysis to the pentose phosphate cycle and to reduced activity of the tricaboxylic acid cycle. As a consequence the production of the prooxidant NADH is reduced and production of the protective NADPH is enhanced. In R. sphaeroides enzymes for glycolysis, pentose phosphate pathway, Entner–Doudoroff pathway and gluconeogenesis are induced in response to 1O2 by the alternative sigma factor RpoHII. The same is true for the sRNA SorY. By limiting malate import SorY thus contributes to the balance of the metabolic fluxes under photooxidative stress conditions. This assigns a so far unknown function to an sRNA in oxidative stress response.
Project description:Distinct retrograde signaling pathways have been identified for several cellular organelles. These pathways are important to maintain the function of these organelles in response to organelle-specific stress. Using Caenorhabditis elegans, we show for the first time that such a retrograde signaling also exists for peroxisomes. Analysis of the C. elegans transcriptome revealed that peroxisomal import stress caused by the knock-down of the peroxisomal matrix protein import receptor prx-5/PEX5 induces the compensatory up-regulation of genes involved in defense response and lipid metabolic processes, especially peroxisomal beta oxidation. We, therefore, propose that the peroxisomal retrograde signaling participates in the maintenance of peroxisomal function in response to peroxisomal import stress.