Browse
Submit Data
Databases
API
Help

Dataset Information

15 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Genetic engineering of AtAOX1a in Saccharomyces cerevisiae prevents oxidative damage and maintains redox homeostasis.

ABSTRACT: This study aimed to validate the physiological importance of Arabidopsis thaliana alternative oxidase 1a (AtAOX1a) in alleviating oxidative stress using Saccharomyces cerevisiae as a model organism. The AOX1a transformant (pYES2AtAOX1a) showed cyanide resistant and salicylhydroxamic acid (SHAM)-sensitive respiration, indicating functional expression of AtAOX1a in S. cerevisiae. After exposure to oxidative stress, pYES2AtAOX1a showed better survival and a decrease in reactive oxygen species (ROS) when compared to S. cerevisiae with empty vector (pYES2). Furthermore, pYES2AtAOX1a sustained growth by regulating GPX2 and/or TSA2, and cellular NAD (+)/NADH ratio. Thus, the expression of AtAOX1a in S. cerevisiae enhances its respiratory tolerance which, in turn, maintains cellular redox homeostasis and protects from oxidative damage.

SUBMITTER: Vishwakarma A

PROVIDER: S-EPMC4821348 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Genetic engineering of AtAOX1a in Saccharomyces cerevisiae prevents oxidative damage and maintains redox homeostasis.

Vishwakarma Abhaypratap A Dalal Ahan A Tetali Sarada Devi SD Kirti Pulugurtha Bharadwaja PB Padmasree Kollipara K

FEBS open bio 20160201 2

This study aimed to validate the physiological importance of Arabidopsis thaliana alternative oxidase 1a (AtAOX1a) in alleviating oxidative stress using Saccharomyces cerevisiae as a model organism. The AOX1a transformant (pYES2AtAOX1a) showed cyanide resistant and salicylhydroxamic acid (SHAM)-sensitive respiration, indicating functional expression of AtAOX1a in S. cerevisiae. After exposure to oxidative stress, pYES2AtAOX1a showed better survival and a decrease in reactive oxygen species (ROS) ...[more]

PMID: 27239435

Similar Datasets

Metabolic engineering of Saccharomyces cerevisiae for astaxanthin production and oxidative stress tolerance.

Project description:The red carotenoid astaxanthin possesses higher antioxidant activity than other carotenoids and has great commercial potential for use in the aquaculture, pharmaceutical, and food industries. In this study, we produced astaxanthin in the budding yeast Saccharomyces cerevisiae by introducing the genes involved in astaxanthin biosynthesis of carotenogenic microorganisms. In particular, expression of genes of the red yeast Xanthophyllomyces dendrorhous encoding phytoene desaturase (crtI product) and bifunctional phytoene synthase/lycopene cyclase (crtYB product) resulted in the accumulation of a small amount of beta-carotene in S. cerevisiae. Overexpression of geranylgeranyl pyrophosphate (GGPP) synthase from S. cerevisiae (the BTS1 gene product) increased the intracellular beta-carotene levels due to the accelerated conversion of farnesyl pyrophosphate to GGPP. Introduction of the X. dendrorhous crtS gene, encoding astaxanthin synthase, assumed to be the cytochrome P450 enzyme, did not lead to astaxanthin production. However, coexpression of CrtS with X. dendrorhous CrtR, a cytochrome P450 reductase, resulted in the accumulation of a small amount of astaxanthin. In addition, the beta-carotene-producing yeast cells transformed by the bacterial genes crtW and crtZ, encoding beta-carotene ketolase and hydroxylase, respectively, also accumulated astaxanthin and its intermediates, echinenone, canthaxanthin, and zeaxanthin. Interestingly, we found that these ketocarotenoids conferred oxidative stress tolerance on S. cerevisiae cells. This metabolic engineering has potential for overproduction of astaxanthin and breeding of novel oxidative stress-tolerant yeast strains.

| S-EPMC2786542 | biostudies-literature

Oxidative damage and mutagenesis in Saccharomyces cerevisiae: genetic studies of pathways affecting replication fidelity of 8-oxoguanine.

Project description:Oxidative damage to DNA constitutes a major threat to the faithful replication of DNA in all organisms and it is therefore important to understand the various mechanisms that are responsible for repair of such damage and the consequences of unrepaired damage. In these experiments, we make use of a reporter system in Saccharomyces cerevisiae that can measure the specific increase of each type of base pair mutation by measuring reversion to a Trp+ phenotype. We demonstrate that increased oxidative damage due to the absence of the superoxide dismutase gene, SOD1, increases all types of base pair mutations and that mismatch repair (MMR) reduces some, but not all, types of mutations. By analyzing various strains that can revert only via a specific CG→AT transversion in backgrounds deficient in Ogg1 (encoding an 8-oxoG glycosylase), we can study mutagenesis due to a known 8-oxoG base. We show as expected that MMR helps prevent mutagenesis due to this damaged base and that Pol η is important for its accurate replication. In addition we find that its accurate replication is facilitated by template switching, as loss of either RAD5 or MMS2 leads to a significant decrease in accurate replication. We observe that these ogg1 strains accumulate revertants during prolonged incubation on plates, in a process most likely due to retromutagenesis.

| S-EPMC3781965 | biostudies-literature

Synthetic Toolkit for Complex Genetic Circuit Engineering in Saccharomyces cerevisiae.

Project description:Sustainable production of chemicals, materials, and pharmaceuticals is increasingly performed by genetically engineered cell factories. Engineering of complex metabolic routes or cell behavior control systems requires robust and predictable gene expression tools. In this challenging task, orthogonality is a fundamental prerequisite for such tools. In this study, we developed and characterized in depth a comprehensive gene expression toolkit that allows accurate control of gene expression in Saccharomyces cerevisiae without marked interference with native cellular regulation. The toolkit comprises a set of transcription factors, designed to function as synthetic activators or repressors, and transcription-factor-dependent promoters, which together provide a broad expression range surpassing, at high end, the strongest native promoters. Modularity of the developed tools is demonstrated by establishing a novel bistable genetic circuit with robust performance to control a heterologous metabolic pathway and enabling on-demand switching between two alternative metabolic branches.

| S-EPMC6150731 | biostudies-literature

Genetic and environmental factors influencing glutathione homeostasis in Saccharomyces cerevisiae.

Project description:Glutathione is an essential metabolite protecting cells against oxidative stress and aging. Here, we show that endogenously synthesized glutathione undergoes intercellular cycling during growth to stationary phase. Genome-wide screening identified approximately 270 yeast deletion mutants that overexcrete glutathione, predominantly in the reduced form, and identified a surprising set of functions important for glutathione homeostasis. The highest excretors were affected in late endosome/vacuolar functions. Other functions identified included nitrogen/carbon source signaling, mitochondrial electron transport, ubiquitin/proteasomal processes, transcriptional regulation, ion transport and the cellular integrity pathway. For many mutants the availability of branched chain amino acids and extracellular pH influenced both glutathione homeostasis and cell viability. For all mutants tested, the onset of glutathione excretion occurred when intracellular concentration exceeded the maximal level found in the parental strain; however, in some mutants prolonged excretion led to substantial depletion of intracellular glutathione. These results significantly contribute to understanding mechanisms affecting glutathione homeostasis in eukaryotes and may provide insight into the underlying cause of glutathione depletion in degenerative processes such as Parkinson's disease. The important implications of these data for use of the yeast deletion collection for the study of other phenomena also are discussed.

| S-EPMC539166 | biostudies-literature

Multistability maintains redox homeostasis in human cells.

Project description:Cells metabolize nutrients through a complex metabolic and signaling network that governs redox homeostasis. At the core of this, redox regulatory network is a mutually inhibitory relationship between reduced glutathione and reactive oxygen species (ROS)-two opposing metabolites that are linked to upstream nutrient metabolic pathways (glucose, cysteine, and glutamine) and downstream feedback loops of signaling pathways (calcium and NADPH oxidase). We developed a nutrient-redox model of human cells to understand system-level properties of this network. Combining in silico modeling and ROS measurements in individual cells, we show that ROS dynamics follow a switch-like, all-or-none response upon glucose deprivation at a threshold that is approximately two orders of magnitude lower than its physiological concentration. We also confirm that this ROS switch can be irreversible and exhibits hysteresis, a hallmark of bistability. Our findings evidence that bistability modulates redox homeostasis in human cells and provide a general framework for quantitative investigations of redox regulation in humans.

| S-EPMC8493564 | biostudies-literature

Altered intracellular calcium homeostasis and endoplasmic reticulum redox state in Saccharomyces cerevisiae cells lacking Grx6 glutaredoxin.

Project description:Glutaredoxin 6 (Grx6) of Saccharomyces cerevisiae is an integral thiol oxidoreductase protein of the endoplasmic reticulum/Golgi vesicles. Its absence alters the redox equilibrium of the reticulum lumen toward a more oxidized state, thus compensating the defects in protein folding/secretion and cell growth caused by low levels of the oxidase Ero1. In addition, null mutants in GRX6 display a more intense unfolded protein response than wild-type cells upon treatment with inducers of this pathway. These observations support a role of Grx6 in regulating the glutathionylation of thiols of endoplasmic reticulum/Golgi target proteins and consequently the equilibrium between reduced and oxidized glutathione in the lumen of these compartments. A specific function influenced by Grx6 activity is the homeostasis of intracellular calcium. Grx6-deficient mutants have reduced levels of calcium in the ER lumen, whereas accumulation occurs at the cytosol from extracellular sources. This results in permanent activation of the calcineurin-dependent pathway in these cells. Some but not all the phenotypes of the mutant are coincident with those of mutants deficient in intracellular calcium transporters, such as the Golgi Pmr1 protein. The results presented in this study provide evidence for redox regulation of calcium homeostasis in yeast cells.

| S-EPMC4279222 | biostudies-literature

Engineering Saccharomyces cerevisiae for production of simvastatin.

Project description:Simvastatin is a semisynthetic cholesterol-lowering medication and one of the top-selling statins in the world. Currently, industrial production of simvastatin acid (SVA) is a multistep process starting from the natural product lovastatin. For this reason, there is significant interest in direct production of simvastatin from a microbial host. In this study, six heterologous biosynthetic genes were introduced into Saccharomyces cerevisiae and the acyl-donor dimethylbutyryl-S-methyl mercaptopropionate (DMB-SMMP) was added, resulting in initial production of 0.5?mg/L SVA. Switching the yeast strain from JHY686 to BJ5464-NpgA increased total polyketide production to over 60?mg/L and conversion from dihydromonacolin L acid to monacolin J acid (MJA) was increased from 60% to 90% by tuning the copy number of the P450 lovA. Increasing the media pH to 8.7 led to a further 10-fold increase in SVA production. Optimized chemical lysis of the cell walls in situ after maximum MJA production led to 55?mg/L SVA titer, representing nearly complete conversion from MJA and a 110-fold increase in titer from the initial SVA production strain. The yeast strains developed in this work can be used as an alternative production method for SVA, and the strategies employed can be broadly applied for heterologous production of other fungal polyketides and semisynthetic compounds in yeast.

| S-EPMC6348118 | biostudies-literature

Nitric oxide maintains endothelial redox homeostasis through PKM2 inhibition

Project description:Decreased nitric oxide (NO) bioavailability and oxidative stress are hallmarks of endothelial dysfunction and cardiovascular diseases. Although numerous proteins are S-nitrosated, whether and how changes in protein S-nitrosation influence endothelial function under pathophysiological conditions remains unknown. We report that active endothelial NO synthase (eNOS) interacts with and S-nitrosates pyruvate kinase M2 (PKM2), which reduces PKM2 activity. PKM2 inhibition increases substrate flux through the pentose phosphate pathway to generate reducing equivalents (NADPH and GSH) and protects against oxidative stress. In mice, the Tyr656 to Phe mutation renders eNOS insensitive to inactivation by oxidative stress and prevents the decrease in PKM2 S-nitrosation and reducing equivalents thereby delaying cardiovascular disease development. These findings highlight a novel mechanism linking NO bioavailability to antioxidant responses in endothelial cells through S-nitrosation and inhibition of PKM2.

| S-SCDT-EMBOJ-2018-100938 | biostudies-other

Chronic oxidative DNA damage due to DNA repair defects causes chromosomal instability in Saccharomyces cerevisiae.

Project description:Oxidative DNA damage is likely to be involved in the etiology of cancer and is thought to accelerate tumorigenesis via increased mutation rates. However, the majority of malignant cells acquire a specific type of genomic instability characterized by large-scale genomic rearrangements, referred to as chromosomal instability (CIN). The molecular mechanisms underlying CIN are not entirely understood. We utilized Saccharomyces cerevisiae as a model system to delineate the relationship between genotoxic stress and CIN. It was found that elevated levels of chronic, unrepaired oxidative DNA damage caused chromosomal aberrations at remarkably high frequencies under both selective and nonselective growth conditions. In this system, exceeding the cellular capacity to appropriately manage oxidative DNA damage resulted in a "gain-of-CIN" phenotype and led to profound karyotypic instability. These results illustrate a novel mechanism for genome destabilization that is likely to be relevant to human carcinogenesis.

| S-EPMC2519736 | biostudies-literature

Decay profiles of Saccharomyces cerevisiae mRNAs following oxidative stress and DNA damage

Project description:We subjected yeast to two stresses, oxidative stress, which under current settings induces a fast and transient response in mRNA abundance, and DNA damage, which triggers a slow enduring response. Using microarrays, we performed a transcriptional arrest experiment to measure genome-wide mRNA decay profiles under each condition. Genome-wide decay kinetics in each condition were compared to decay experiments that were performed in a reference condition (only transcription inhibition without an additional stress) to quantify changes in mRNA stability in each condition. We found condition-specific changes in mRNA decay rates and coordination between mRNA production and degradation. In the transient response, most induced genes were surprisingly destabilized, while repressed genes were somewhat stabilized, exhibiting counteraction between production and degradation. This strategy can reconcile high steady-state level with short response time among induced genes. In contrast, the stress that induces the slow response displays the more expected behavior, whereby most induced genes are stabilized, and repressed genes destabilized. Our results show genome-wide interplay between mRNA production and degradation, and that alternative modes of such interplay determine the kinetics of the transcriptome in response to stress. Keywords: Four separate time courses

2008-10-01 | GSE12221 | GEO

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data