scholarly journals Monitoring yeast mitochondria with peroxiredoxin-based redox probes: the influence of oxygen and glucose availability

2017 ◽  
Vol 7 (2) ◽  
pp. 20160143 ◽  
Author(s):  
Daniel Pastor-Flores ◽  
Katja Becker ◽  
Tobias P. Dick
Keyword(s):  
Pentose Phosphate Pathway ◽  
Redox State ◽  
Pentose Phosphate ◽  
Yeast Cells ◽  
Mitochondrial Matrix ◽  
Yeast Mitochondria ◽  
Dominant Source ◽  
Significant Interest ◽  
Metabolic Conditions ◽  
Glucose Availability

Mitochondrially generated oxidants are believed to play important roles in both physiology and pathophysiology. Therefore, it is of significant interest to better understand the metabolic conditions leading to enhanced mitochondrial oxidant generation. Here, we investigate the influence of oxygen and glucose availability on the redox state of peroxiredoxin-based redox probes, expressed in the cytosol and mitochondrial matrix of yeast cells. We observe that the redox state of peroxiredoxin probes reflects the balance between dioxygen-dependent peroxide generation and glucose-dependent generation of reducing equivalents. The oxidative pentose phosphate pathway appears to be the dominant source of NADPH in the system under study.

Hyperpolarized [U-2H, U-13C]Glucose reports on glycolytic and pentose phosphate pathway activity in EL4 tumors and glycolytic activity in yeast cells

2014 ◽  
Vol 74 (6) ◽  
pp. 1543-1547 ◽  
Author(s):  
Kerstin N. Timm ◽  
Johannes Hartl ◽  
Markus A. Keller ◽  
De-En Hu ◽  
Mikko I. Kettunen ◽  
...  
Keyword(s):  
Pentose Phosphate Pathway ◽  
Pentose Phosphate ◽  
Yeast Cells ◽  
Pathway Activity ◽  
Glycolytic Activity

scholarly journals Novel Proteomic changes in Yeast Mitochondria provide insights into mitochondrial functioning upon over-expression of human p53

2019 ◽  
Author(s):  
Archana Kumari Redhu ◽  
Jayadeva Paike Bhat
Keyword(s):  
Carbon Source ◽  
Warburg Effect ◽  
Pentose Phosphate ◽  
Yeast Cells ◽  
Yeast Mitochondria ◽  
Over Expression ◽  
Cycle Arrest ◽  
Glycolytic Activity ◽  
Sucrose Fermentation ◽  
Induced Apoptosis

AbstractCancer cells display enhanced glycolytic activity and impaired oxidative phosphorylation even in the presence of adequate oxygen (Warburg effect). Mitochondrial physiology is a promising hit target for anti-cancer therapy because of its key role in Warburg effect and activating apoptosis in mammalian as well as yeast cells. Over-expression of human p53 in S.cerevisiae leads to cell cycle arrest and apotosis. In the present work we show that how S.cerevisiae escapes from p53 induced apoptosis in fermentable carbon source, whereas in case of non-fermentable carbon source this phenomenon is not observed. To shed the light on this aspect we performed a quantitative proteomic analysis of yeast mitochondria isolated from the cells grown on sucrose (fermentation) and glycerol (respiration) with and without p53 over-expression. Through this approach, we identified a total dataset of 1120 proteins with 1% FDR, of which 239(133+106) proteins are differentially experssed in both conditions. Interestingly, we observed that after over-expression of p53 in sucrose grown yeast cells, a complete set of pentose phosphate pathway (PPP) enzymes is up-regulated in the mitochondria that leads to enhanced mitochondrial NADPH production and ROS quenching. Increased association of a hexose transporter (HXT6) and a hexokinase (HXK2) with the mitochondria of fermenting yeast cells upon over-expression of p53, may direct glucose towards PPP inside the mitochondria. In conclusion, our results provide the evidence that up-regulated PPP inside the mitochondria is a key to evade apoptosis by S.cerevisiae upon p53 over-expression.

scholarly journals Protein S-thiolation targets glycolysis and protein synthesis in response to oxidative stress in the yeast Saccharomyces cerevisiae

2003 ◽  
Vol 374 (2) ◽  
pp. 513-519 ◽  
Author(s):  
Daniel SHENTON ◽  
Chris M. GRANT
Keyword(s):  
Oxidative Stress ◽  
Protein Synthesis ◽  
Alcohol Dehydrogenase ◽  
Pentose Phosphate Pathway ◽  
Protein S ◽  
Pentose Phosphate ◽  
Yeast Cells ◽  
Glycolytic Flux ◽  
Irreversible Oxidation ◽  
Peroxide Stress

The irreversible oxidation of cysteine residues can be prevented by protein S-thiolation, a process by which protein SH groups form mixed disulphides with low-molecular-mass thiols such as glutathione. We report here the target proteins which are modified in yeast cells in response to H2O2. In particular, a range of glycolytic and related enzymes (Tdh3, Eno2, Adh1, Tpi1, Ald6 and Fba1), as well as translation factors (Tef2, Tef5, Nip1 and Rps5) are identified. The oxidative stress conditions used to induce S-thiolation are shown to inhibit GAPDH (glyceraldehyde-3-phosphate dehydrogenase), enolase and alcohol dehydrogenase activities, whereas they have no effect on aldolase, triose phosphate isomerase or aldehyde dehydrogenase activities. The inhibition of GAPDH, enolase and alcohol dehydrogenase is readily reversible once the oxidant is removed. In addition, we show that peroxide stress has little or no effect on glucose-6-phosphate dehydrogenase or 6-phosphogluconate dehydrogenase, the enzymes that catalyse NADPH production via the pentose phosphate pathway. Thus the inhibition of glycolytic flux is proposed to result in glucose equivalents entering the pentose phosphate pathway for the generation of NADPH. Radiolabelling is used to confirm that peroxide stress results in a rapid and reversible inhibition of protein synthesis. Furthermore, we show that glycolytic enzyme activities and protein synthesis are irreversibly inhibited in a mutant that lacks glutathione, and hence cannot modify proteins by S-thiolation. In summary, protein S-thiolation appears to serve an adaptive function during exposure to an oxidative stress by reprogramming metabolism and protecting protein synthesis against irreversible oxidation.

scholarly journals Systematic Identification of Regulators of Oxidative Stress Reveals Non-canonical Roles for Peroxisomal Import and the Pentose Phosphate Pathway

Cell Reports ◽  
2020 ◽  
Vol 30 (5) ◽  
pp. 1417-1433.e7 ◽  
Author(s):  
Michael M. Dubreuil ◽  
David W. Morgens ◽  
Kanji Okumoto ◽  
Masanori Honsho ◽  
Kévin Contrepois ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Pentose Phosphate Pathway ◽  
Pentose Phosphate ◽  
Systematic Identification

scholarly journals Geranylgeranylacetone attenuates cerebral ischemia–reperfusion injury in rats through the augmentation of HSP 27 phosphorylation: a preliminary study

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Kazuya Matsuo ◽  
Kohkichi Hosoda ◽  
Jun Tanaka ◽  
Yusuke Yamamoto ◽  
Taichiro Imahori ◽  
...  
Keyword(s):  
Cerebral Ischemia ◽  
Reperfusion Injury ◽  
Pentose Phosphate Pathway ◽  
Ischemia Reperfusion Injury ◽  
Ischemia Reperfusion ◽  
Pentose Phosphate ◽  
Cerebral Ischemia Reperfusion ◽  
Hsp27 Phosphorylation ◽  
Cerebral Ischemia Reperfusion Injury ◽  
Preliminary Study

Abstract Background We previously reported that heat shock protein 27 (HSP27) phosphorylation plays an important role in the activation of glucose-6-phosphate dehydrogenase (G6PD), resulting in the upregulation of the pentose phosphate pathway and antioxidant effects against cerebral ischemia–reperfusion injury. The present study investigated the effect of geranylgeranylacetone, an inducer of HSP27, on ischemia–reperfusion injury in male rats as a preliminary study to see if further research of the effects of geranylgeranylacetone on the ischemic stroke was warranted. Methods In all experiments, male Wistar rats were used. First, we conducted pathway activity profiling based on a gas chromatography–mass spectrometry to identify ischemia–reperfusion-related metabolic pathways. Next, we investigated the effects of geranylgeranylacetone on the pentose phosphate pathway and ischemia–reperfusion injury by real-time polymerase chain reaction (RT-PCR), immunoblotting, and G6PD activity, protein carbonylation and infarct volume analysis. Geranylgeranylacetone or vehicle was injected intracerebroventricularly 3 h prior to middle cerebral artery occlusion or sham operation. Results Pathway activity profiling demonstrated that changes in the metabolic state depended on reperfusion time and that the pentose phosphate pathway and taurine-hypotaurine metabolism pathway were the most strongly related to reperfusion among 137 metabolic pathways. RT-PCR demonstrated that geranylgeranylacetone did not significantly affect the increase in HSP27 transcript levels after ischemia–reperfusion. Immunoblotting showed that geranylgeranylacetone did not significantly affect the elevation of HSP27 protein levels. However, geranylgeranylacetone significantly increase the elevation of phosphorylation of HSP27 after ischemia–reperfusion. In addition, geranylgeranylacetone significantly affected the increase in G6PD activity, and reduced the increase in protein carbonylation after ischemia–reperfusion. Accordingly, geranylgeranylacetone significantly reduced the infarct size (median 31.3% vs 19.9%, p = 0.0013). Conclusions As a preliminary study, these findings suggest that geranylgeranylacetone may be a promising agent for the treatment of ischemic stroke and would be worthy of further study. Further studies are required to clearly delineate the mechanism of geranylgeranylacetone-induced HSP27 phosphorylation in antioxidant effects, which may guide the development of new approaches for minimizing the impact of cerebral ischemia–reperfusion injury.

scholarly journals Pentose Phosphate Pathway Reactions in Photosynthesizing Cells

Cells ◽  
2021 ◽  
Vol 10 (6) ◽  
pp. 1547
Author(s):  
Thomas D. Sharkey
Keyword(s):  
Pentose Phosphate Pathway ◽  
Strong Association ◽  
Pentose Phosphate ◽  
Critical Component ◽  
Reaction Sequence ◽  
Response To Stress ◽  
Labeling Pattern

The pentose phosphate pathway (PPP) is divided into an oxidative branch that makes pentose phosphates and a non-oxidative branch that consumes pentose phosphates, though the non-oxidative branch is considered reversible. A modified version of the non-oxidative branch is a critical component of the Calvin–Benson cycle that converts CO2 into sugar. The reaction sequence in the Calvin–Benson cycle is from triose phosphates to pentose phosphates, the opposite of the typical direction of the non-oxidative PPP. The photosynthetic direction is favored by replacing the transaldolase step of the normal non-oxidative PPP with a second aldolase reaction plus sedoheptulose-1,7-bisphosphatase. This can be considered an anabolic version of the non-oxidative PPP and is found in a few situations other than photosynthesis. In addition to the strong association of the non-oxidative PPP with photosynthesis metabolism, there is recent evidence that the oxidative PPP reactions are also important in photosynthesizing cells. These reactions can form a shunt around the non-oxidative PPP section of the Calvin–Benson cycle, consuming three ATP per glucose 6-phosphate consumed. A constitutive operation of this shunt occurs in the cytosol and gives rise to an unusual labeling pattern of photosynthetic metabolites while an inducible shunt in the stroma may occur in response to stress.

scholarly journals Fine tuning the glycolytic flux ratio of EP-bifido pathway for mevalonate production by enhancing glucose-6-phosphate dehydrogenase (Zwf) and CRISPRi suppressing 6-phosphofructose kinase (PfkA) in Escherichia coli

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Ying Li ◽  
He Xian ◽  
Ya Xu ◽  
Yuan Zhu ◽  
Zhijie Sun ◽  
...  
Keyword(s):  
Pentose Phosphate Pathway ◽  
Conversion Rate ◽  
Metabolic Flux ◽  
Flux Ratio ◽  
Reducing Power ◽  
Pentose Phosphate ◽  
Fine Tuning ◽  
High Yield ◽  
Glycolytic Flux ◽  
Acetyl Coa

Abstract Background Natural glycolysis encounters the decarboxylation of glucose partial oxidation product pyruvate into acetyl-CoA, where one-third of the carbon is lost at CO2. We previously constructed a carbon saving pathway, EP-bifido pathway by combining Embden-Meyerhof-Parnas Pathway, Pentose Phosphate Pathway and “bifid shunt”, to generate high yield acetyl-CoA from glucose. However, the carbon conversion rate and reducing power of this pathway was not optimal, the flux ratio of EMP pathway and pentose phosphate pathway (PPP) needs to be precisely and dynamically adjusted to improve the production of mevalonate (MVA). Result Here, we finely tuned the glycolytic flux ratio in two ways. First, we enhanced PPP flux for NADPH supply by replacing the promoter of zwf on the genome with a set of different strength promoters. Compared with the previous EP-bifido strains, the zwf-modified strains showed obvious differences in NADPH, NADH, and ATP synthesis levels. Among them, strain BP10BF accumulated 11.2 g/L of MVA after 72 h of fermentation and the molar conversion rate from glucose reached 62.2%. Second, pfkA was finely down-regulated by the clustered regularly interspaced short palindromic repeats interference (CRISPRi) system. The MVA yield of the regulated strain BiB1F was 8.53 g/L, and the conversion rate from glucose reached 68.7%. Conclusion This is the highest MVA conversion rate reported in shaken flask fermentation. The CRISPRi and promoter fine-tuning provided an effective strategy for metabolic flux redistribution in many metabolic pathways and promotes the chemicals production.

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