Trx2p-dependent regulation of Saccharomyces cerevisiae oxidative stress response by the Skn7p transcription factor under respiring conditions

The whole genome analysis has demonstrated that wine yeasts undergo changes in promoter regions and variations in gene copy number, which make them different to lab strains and help them better adapt to stressful conditions during winemaking, where oxidative stress plays a critical role. Since cytoplasmic thioredoxin II, a small protein with thiol-disulphide oxidoreductase activity, has been seen to perform important functions under biomass propagation conditions of wine yeasts, we studied the involvement of Trx2p in the molecular regulation of the oxidative stress transcriptional response on these strains. In this study, we analyzed the expression levels of several oxidative stress-related genes regulated by either Yap1p or the co-operation between Yap1p and Skn7p. The results revealed a lowered expression for all the tested Skn7p dependent genes in a Trx2p-deficient strain and that Trx2p is essential for the oxidative stress response during respiratory metabolism in wine yeast. Additionally, activity of Yap1p and Skn7p dependent promoters by β-galactosidase assays clearly demonstrated that Skn7p-dependent promoter activation is affected by TRX2 gene deficiency. Finally we showed that deleting the TRX2 gene causes Skn7p hyperphosphorylation under oxidative stress conditions. We propose Trx2p to be a new positive efector in the regulation of the Skn7p transcription factor that controls phosphorylation events and, therefore, modulates the oxidative stress response in yeast. © 2013 Gómez-Pastor et al.This work has been supported by grants AGL 2008-00060 and AGL2011-24353 from the Spanish Ministry of Education and Science (MEC) to EM. EG was a predoctoral fellow of the FPI program from the MEC (Spanish Ministry of Education). RG-P was a predoctoral fellow of the I3P program from the CSIC (Spanish National Research Council). RP-T was a postdoctoral fellow of the JAEDOC program from the CSIC.Peer Reviewe

Reduction of oxidative cellular damage by overexpression of the thioredoxin TRX2 gene improves yield and quality of wine yeast dry active biomass

<p>Abstract</p> <p>Background</p> <p>Wine <it>Saccharomyces cerevisiae </it>strains, adapted to anaerobic must fermentations, suffer oxidative stress when they are grown under aerobic conditions for biomass propagation in the industrial process of active dry yeast production. Oxidative metabolism of sugars favors high biomass yields but also causes increased oxidation damage of cell components. The overexpression of the <it>TRX2 </it>gene, coding for a thioredoxin, enhances oxidative stress resistance in a wine yeast strain model. The thioredoxin and also the glutathione/glutaredoxin system constitute the most important defense against oxidation. Trx2p is also involved in the regulation of Yap1p-driven transcriptional response against some reactive oxygen species.</p> <p>Results</p> <p>Laboratory scale simulations of the industrial active dry biomass production process demonstrate that <it>TRX2 </it>overexpression increases the wine yeast final biomass yield and also its fermentative capacity both after the batch and fed-batch phases. Microvinifications carried out with the modified strain show a fast start phenotype derived from its enhanced fermentative capacity and also increased content of beneficial aroma compounds. The modified strain displays an increased transcriptional response of Yap1p regulated genes and other oxidative stress related genes. Activities of antioxidant enzymes like Sod1p, Sod2p and catalase are also enhanced. Consequently, diminished oxidation of lipids and proteins is observed in the modified strain, which can explain the improved performance of the thioredoxin overexpressing strain.</p> <p>Conclusions</p> <p>We report several beneficial effects of overexpressing the thioredoxin gene <it>TRX2 </it>in a wine yeast strain. We show that this strain presents an enhanced redox defense. Increased yield of biomass production process in <it>TRX2 </it>overexpressing strain can be of special interest for several industrial applications.</p

Correction: reduction of oxidative cellular damage by overexpression of the thioredoxin TRX2 gene improves yield and quality of wine yeast dry active biomass

AbstractFollowing publication of this work [Gomez-Pastor et al, Microbial Cell Factories 2010, 9:9] we have noticed a production error in the article. Figure 1 in the original version showed incorrect results, with graphs having been duplicated in error from another figure. The correct results for Figure 1 are shown below. Legend to Figure 1 Improved performance of TTRX2 strain in biomass production process. (A) Biomass produced (continuous line) and oxygen saturation (discontinuous line) along bench-top trials of biomass propagation for T73 (black diamond), TTRX2 (white square) and TGSH1 (white triangle) strains by measuring OD600 from diluted samples. Average of three independent experiments and standard deviations are shown. (B) Fermentative capacity of yeast biomass collected at the end of the batch and fed-batch stages of growth in bench-top trials of ADY production. Biomass from wild-type T73 (black bars) and TTRX2 (white bars) were dehydrated until 8% moisture before performing the analysis. Data were normalized to the fermentative capacity of the batch sample from T73 strain. Average of three independent experiments and standard deviations are shown. Significantly different values compared to the control (p < 0.001) were marked by asterisk. (C) Sugar consumption profiles during microvinification experiments using natural Bobal must for T73 (closed symbols) and TTRX2 (open symbols) strains. The start of must fermentation was followed in detail during the first 6 hours for both strains T73 (closed symbol) and TTRX2 (open symbol). Averages were obtained from two independent experiments with three technical replicates for each one.Peer Reviewe

Engineered Trx2p industrial yeast strain protects glycolysis and fermentation proteins from oxidative carbonylation during biomass propagation

<p>Abstract</p> <p>Background</p> <p>In the yeast biomass production process, protein carbonylation has severe adverse effects since it diminishes biomass yield and profitability of industrial production plants. However, this significant detriment of yeast performance can be alleviated by increasing thioredoxins levels. Thioredoxins are important antioxidant defenses implicated in many functions in cells, and their primordial functions include scavenging of reactive oxygen species that produce dramatic and irreversible alterations such as protein carbonylation.</p> <p>Results</p> <p>In this work we have found several proteins specifically protected by yeast Thioredoxin 2 (Trx2p). Bidimensional electrophoresis and carbonylated protein identification from <it>TRX</it>-deficient and <it>TRX</it>-overexpressing cells revealed that glycolysis and fermentation-related proteins are specific targets of Trx2p protection. Indeed, the <it>TRX2 </it>overexpressing strain presented increased activity of the central carbon metabolism enzymes. Interestingly, Trx2p specifically preserved alcohol dehydrogenase I (Adh1p) from carbonylation, decreased oligomer aggregates and increased its enzymatic activity.</p> <p>Conclusions</p> <p>The identified proteins suggest that the fermentative capacity detriment observed under industrial conditions in T73 wine commercial strain results from the oxidative carbonylation of specific glycolytic and fermentation enzymes. Indeed, increased thioredoxin levels enhance the performance of key fermentation enzymes such as Adh1p, which consequently increases fermentative capacity.</p

Effects of pharmacological agents on the lifespan phenotype of Drosophila DJ-1β mutants

Mutations in the DJ-1 gene cause autosomal recessive, early-onset Parkinsonism. The DJ-1 protein exerts a protective role against oxidative stress damage, working as a cellular oxidative stress sensor, and it seems to regulate gene expression at different levels. In Drosophila, two DJ-1 orthologs have been identified: DJ-1β and DJ-1β. Several studies have shown that loss of DJ-1β function causes Parkinson's disease (PD)-like phenotypes in flies such as age-dependent locomotor defects, reduced lifespan, and enhanced sensitivity to toxins that induce oxidative stress, like the herbicide paraquat. However, no dopaminergic neurodegeneration is observed. These results suggested that both locomotor and lifespan phenotypes could be either related to defects in oxidative stress response, or in dopaminergic physiology as proposed in mice models. In this study, we have employed pharmacological approaches to modify the lifespan phenotype of DJ-1β mutant flies. We have assessed the effects of chronic treatments with antiparkinsonian drugs as well as with antioxidant compounds on such phenotype finding that only antioxidants show statistically significant beneficial effects on DJ-1β mutants' lifespan. These results strongly suggest that oxidative stress plays a causal role in the lifespan phenotype of DJ-1β mutants. Consistent with this, we find that loss of DJ-1β function results in cellular accumulation of reactive oxygen species (ROS) in adult brains, elevated levels of lipid peroxidation and an increased catalase enzymatic activity, thus indicating the existence of high oxidative stress levels in DJ-1β mutants and confirming the essential function of the DJ-1β protein in protecting the organism against oxidative insults. Our study further shows that the lifespan phenotype of DJ-1β mutant flies is amenable to pharmacological intervention, and validates Drosophila as a valuable model for testing and identifying new drugs with therapeutic potential for PD. © 2010 Elsevier B.V.E. L.-C. was supported by a predoctoral fellowship from Consellería de Cultura, Educació i Ciència and R.G.-P. by a predoctoral fellowship of the I3P program from Consejo Superior de Investigaciones Científicas. V. M.-S. is a postdoctoral researcher funded by the Ministerio de Educación y Ciencia. This work has been supported by grants from Consellería de Cultura, Educació i Ciència and, in part, by grants from the Ministerio de Educación y Ciencia to N.P.Peer Reviewe

Non-canonical regulation of glutathione and trehalose biosynthesis characterizes non-Saccharomyces wine yeasts with poor performance in active dry yeast production

Several yeast species, belonging to Saccharomyces and non-Saccharomyces genera, play fundamental roles during spontaneous must grape fermentation, and recent studies have shown that mixed fermenta-tions, co-inoculated with S. cerevisiae and non-Saccharomyces strains, can improve wine organoleptic properties. During active dry yeast (ADY) produc-tion, antioxidant systems play an essential role in yeast survival and vitality as both biomass propagation and dehydration cause cellular oxidative stress and negatively affect technological performance. Mechanisms for adaptation and resistance to desiccation have been described for S. cerevisiae, but no data are available on the physiology and oxidative stress response of non-Saccharomyces wine yeasts and their potential impact on ADY production. In this study we analyzed the oxidative stress response in several non-Saccharomyces yeast species by measuring the activity of reactive oxygen species (ROS) scavenging enzymes, e.g., catalase and glutathione reductase, accumulation of protective metabolites, e.g., trehalose and reduced glutathi-one (GSH), and lipid and protein oxidation levels. Our data suggest that non-canonical regulation of glutathione and trehalose biosynthesis could cause poor fermentative performance after ADY production, as it corroborates the corrective effect of antioxidant treatments, during biomass propagation, with both pure chemicals and food-grade argan oil

LPAC syndrome associated with deletion of the full exon 4 in a ABCB4 genetic mutation in a patient with hepatitis C

El síndrome LPAC (low-phospholipid-associated cholelithiasis syndrome) está asociado a mutaciones del gen ABCB4, que codifica la proteína MDR3, esencial en la secreción de fosfatidilcolina en las sales biliares. Este síndrome se caracteriza por una mayor prevalencia en mujeres, síntomas biliares en adultos jóvenes y excelente respuesta al ácido ursodesoxicólico (AUDC). Presentamos el caso de un hombre de 48 años con hepatitis C, genotipo 1b, fibrosis F3, nula respuesta Peg-IFN-α-2b/ribavirina y cólicos nefríticos de repetición. En 2011 desarrolló ictericia, prurito y dolor cólico epigástrico acompañado de aumento sérico de AST, ALT, GGT, bilirrubina y alfafetoproteína, y carga viral (14.600.000 UI/ml). La endoscopia oral, la ecoendoscopia, la angio-TAC y la ecografía-doppler evidenciaron hepatopatía crónica no cirrótica. El cuadro se autolimitó y un año después sufrió un episodio similar. Iniciamos tratamiento con AUDC, con excelente respuesta clínica. El estudio inmunohistoquímico y la secuenciación completa del gen ABCB4 no mostraron alteraciones. La técnica MLPA® detectó deleción heterocigota del exón 4 completo y confirmó la sospecha de síndrome LPAC.Low-phospholipid-associated cholelithiasis syndrome (LPAC) is associated with ABCB4 genetic mutation. ABCB4 encodes MDR3 protein, involved in biliary phosphatidylcholine excretion. Higher prevalence in women, biliary symptoms in young adults and ursodesoxycholic acid (UDCA) response are the main features. We report the case of a 48-year-old man with hepatitis C, genotype 1b, fibrosis F3, null responder to Peg-IFNα2b/ribavirin and nephritic colic. In 2011 he developed jaundice, pruritus and epigastric pain. He showed increased serum levels of AST, ALT, GGT, bilirubin and alpha-fetoprotein, and viral load (14,600,000IU/mL). Pancreatic CT, endoscopic ultrasonography and echo-Doppler showed non cirrhotic chronic liver disease. The episode resolved spontaneously and one year later he suffered a similar episode. UDCA was started with excellent response. An immunohistochemistry study and sequencing of ABCB4 did not find alteration. MLPA® technique detected heterozygous deletion of the full exon 4 confirming LPAC syndrome diagnosis

Liver injury in non-alcoholic fatty liver disease is associated with urea cycle enzyme dysregulation

The main aim was to evaluate changes in urea cycle enzymes in NAFLD patients and in two preclinical animal models mimicking this entity. Seventeen liver specimens from NAFLD patients were included for immunohistochemistry and gene expression analyses. Three-hundred-and-eighty-two biopsy-proven NAFLD patients were genotyped for rs1047891, a functional variant located in carbamoyl phosphate synthetase-1 (CPS1) gene. Two preclinical models were employed to analyse CPS1 by immunohistochemistry, a choline deficient high-fat diet model (CDA-HFD) and a high fat diet LDLr knockout model (LDLr −/−). A significant downregulation in mRNA was observed in CPS1 and ornithine transcarbamylase (OTC1) in simple steatosis and NASH-fibrosis patients versus controls. Further, age, obesity (BMI > 30 kg/m), diabetes mellitus and ALT werefound to be risk factors whereas A-allele from CPS1 was a protective factor from liver fibrosis. CPS1 hepatic expression was diminished in parallel with the increase of fibrosis, and its levels reverted up to normality after changing diet in CDA-HFD mice. In conclusion, liver fibrosis and steatosis were associated with a reduction in both gene and protein expression patterns of mitochondrial urea cycle enzymes. A-allele from a variant on CPS1 may protect from fibrosis development. CPS1 expression is restored in a preclinical model when the main trigger of the liver damage disappears.The research leading to these results has received funding from the Consejería de Salud de la Junta de Andalucía under grant agreement PC-0148-2016-0148 and PE-0451-2018 and Instituto de Salud Carlos III under grant agreements CD21/00095, PI16/01842, PI19/01404, PI19/00589, IFI18/00041, FI20/00201, CD18/00126 and EHD18PI04/2021. Rocío Gallego-Durán has received the Andrew K Burroughs Fellowship from European Association for the Study of the Liver (EASL), Aprendizaje de Nuevas Tecnologías fellowship from Asociación Española para el Estudio del Hígado (AEEH) and CIBERehd Grant to support researcher’s mobility

Neurodegeneration, Mitochondria, and Antibiotics

Neurodegenerative diseases are characterized by the progressive loss of neurons, synapses, dendrites, and myelin in the central and/or peripheral nervous system. Actual therapeutic options for patients are scarce and merely palliative. Although they affect millions of patients worldwide, the molecular mechanisms underlying these conditions remain unclear. Mitochondrial dysfunction is generally found in neurodegenerative diseases and is believed to be involved in the pathomechanisms of these disorders. Therefore, therapies aiming to improve mitochondrial function are promising approaches for neurodegeneration. Although mitochondrial-targeted treatments are limited, new research findings have unraveled the therapeutic potential of several groups of antibiotics. These drugs possess pleiotropic effects beyond their anti-microbial activity, such as anti-inflammatory or mitochondrial enhancer function. In this review, we will discuss the controversial use of antibiotics as potential therapies in neurodegenerative diseases.This project was supported by FIS PI19/00377 (2019) and FIS PI22/00142 (2022) grants, Instituto de Salud Carlos III, Spain; and the Fondo Europeo de Desarrollo Regional (FEDER Unión Europea), Spanish Ministry of Education, Culture, and Sport. This activity was co-financed by the European Regional Development Fund (ERDF) and by the Regional Ministry of Economic Transformation, Industry, Knowledge, and Universities of the Junta de Andalucía, within the framework of the ERDF Andalusia operational program 2014–2020 Thematic objective “01—Reinforcement of research, technological development and innovation” through the reference research project CTS-5725 and PY18-850.Peer reviewe