Global expression studies in baker's yeast reveal target genes for the improvement of industrially-relevant traits: the cases of CAF16 and ORC2

<p>Abstract</p> <p>Background</p> <p>Recent years have seen a huge growth in the market of industrial yeasts with the need for strains affording better performance or to be used in new applications. Stress tolerance of commercial <it>Saccharomyces cerevisiae </it>yeasts is, without doubt, a trait that needs improving. Such trait is, however, complex, and therefore only in-depth knowledge of their biochemical, physiological and genetic principles can help us to define improvement strategies and to identify the key factors for strain selection.</p> <p>Results</p> <p>We have determined the transcriptional response of commercial baker's yeast cells to both high-sucrose and lean dough by using DNA macroarrays and liquid dough (LD) model system. Cells from compressed yeast blocks display a reciprocal transcription program to that commonly reported for laboratory strains exposed to osmotic stress. This discrepancy likely reflects differences in strain background and/or experimental design. Quite remarkably, we also found that the transcriptional response of starved baker's yeast cells was qualitatively similar in the presence or absence of sucrose in the LD. Nevertheless, there was a set of differentially regulated genes, which might be relevant for cells to adapt to high osmolarity. Consistent with this, overexpression of <it>CAF16 </it>or <it>ORC2</it>, two transcriptional factor-encoding genes included in this group, had positive effects on leavening activity of baker's yeast. Moreover, these effects were more pronounced during freezing and frozen storage of high-sucrose LD.</p> <p>Conclusions</p> <p>Engineering of differentially regulated genes opens the possibility to improve the physiological behavior of baker's yeast cells under stress conditions like those encountered in downstream applications.</p

The oxidation of trichloroethylene over different mixed oxides derived from hydrotalcites

[EN] The activity of different Mg(Fe/Al), Ni(Fe/Al) and Co(Fe/Al) mixed oxides based on hydrotalcite-like compounds have been studied for the catalytic oxidation of trichloroethylene. It has been shown that the Co catalysts are more active than the Ni catalyst, being the Mg catalysts the less active ones. The activity of all the catalysts improves when iron is substituted by aluminum in the catalyst composition. The best results have been obtained with the CoAl mixed oxide derived from hydrotalcite that is a stable, highly active and selective catalyst. These results have been related with the presence of aluminum in the Co3O4 structure that favors, in the presence of oxygen, the formation of O2 − sites and enhances the acid properties of the catalyst. The combination of both characteristics maximizes the adsorption and oxidation of the TCE.The authors wish to thank financial support from CONACYT (project 154060) and from the Spanish Ministry of Economy and Competitiveness through the Consolider Ingenio Multicat (CSD-2009-00050) and MAT-2012-38567-C02-01 programms. N.B.R. acknowledges Catedra Cemex Sostenibilidad (UPV) for a fellowship.Blanch Raga, N.; Palomares Gimeno, AE.; Martínez Triguero, LJ.; Puche Panadero, M.; Fetter, G.; Bosch, P. (2014). The oxidation of trichloroethylene over different mixed oxides derived from hydrotalcites. Applied Catalysis B: Environmental. 160-61:129-134. https://doi.org/10.1016/j.apcatb.2014.05.014129134160-6

Metabolic reprogramming by Acly inhibition using SB-204990 alters glucoregulation and modulates molecular mechanisms associated with aging

19 Páginas.-- 7 FigurasATP-citrate lyase is a central integrator of cellular metabolism in the interface of protein, carbohydrate, and lipid metabolism. The physiological consequences as well as the molecular mechanisms orchestrating the response to long-term pharmacologically induced Acly inhibition are unknown. We report here that the Acly inhibitor SB-204990 improves metabolic health and physical strength in wild-type mice when fed with a high-fat diet, while in mice fed with healthy diet results in metabolic imbalance and moderated insulin resistance. By applying a multiomic approach using untargeted metabolomics, transcriptomics, and proteomics, we determined that, in vivo, SB-204990 plays a role in the regulation of molecular mechanisms associated with aging, such as energy metabolism, mitochondrial function, mTOR signaling, and folate cycle, while global alterations on histone acetylation are absent. Our findings indicate a mechanism for regulating molecular pathways of aging that prevents the development of metabolic abnormalities associated with unhealthy dieting. This strategy might be explored for devising therapeutic approaches to prevent metabolic diseases.This work was funded by grants from the Ministerio de Economía y Competitividad, Instituto de Salud Carlos III, co-funded by Fondos FEDER (PI15/00134, PI18/01590, CPII19/00023 to A.M.M.) and the Ministerio de Ciencia e Innovación (PID2021-123965OB-100 to A.M.M.). A.M.M. is funded by the Junta de Andalucía P20_00480, the Spanish Society of Diabetes, and CSIC 202220I059. M.S.K. is funded by the Nordea Foundation (#02-2017-1749), the Novo Nordisk Foundation (#NNF17OC0027812), the Neye Foundation, the Lundbeck Foundation (#R324-2019-1492), the Ministry of Higher Education and Science of Denmark (#0238-00003B). V.C.G. is funded by the Instituto de Salud Carlos III (CP19/00046), co-funded by FEDER. F.M. is funded by the CIBERDEM of the Instituto de Salud Carlos III. A.M.M. is the guarantor of this work and, as such, had full access to all the data in the study and takes responsibility for the integrity of the data and the accuracy of the data analysis. We acknowledge the support of the group of basic research on diabetes of the Spanish Society of Diabetes.Peer reviewe

Heterologous expression of type I antifreeze peptide GS-5 in baker's yeast increases freeze tolerance and provides enhanced gas production in frozen dough

The demand for frozen-dough products has increased notably in the baking industry. Nowadays, no appropriate industrial baker's yeast with optimal gassing capacity in frozen dough is, however, available, and it is unlikely that classical breeding programs could provide significant improvements of this trait. Antifreeze proteins, found in diverse organisms, display the ability to inhibit the growth of ice, allowing them to survive at temperatures below 0°C. In this study a recombinant antifreeze peptide GS-5 was expressed from tne polar fish grubby sculpin (Myoxocephalus aenaeus) in laboratory and industrial baker's yeast strains of Saccharomyces cerevisiae. Production of the recombinant protein increased freezing tolerance in both strains tested. Furthermore, expression of the GS-5 encoding gene enhanced notably the gassing rate and total gas production in frozen and frozen sweet doughs. These effects are unlikely to be due to reduced osmotic damage during freezing/ thawing, because recombinant cells showed growth behavior similar to that of the parent under hypermosmotic stress conditions.This research was supported by the Comisión Interministerial de Ciencia y Tecnología project (PACTI, COO1999AX173) from the Ministry of Science and Technology of Spain. J.P. is supported by a F.P.I. fellowship.Peer Reviewe

Validation of a Flour-Free Model Dough System for Throughput Studies of Baker's Yeast

Evaluation of gene expression in baker's yeast requires the extraction and collection of pure samples of RNA. However, in bread dough this task is difficult due to the complex composition of the system. We found that a liquid model system can be used to analyze the transcriptional response of industrial strains in dough with a high sugar content. The production levels of CO2 and glycerol by two commercial strains in liquid and flour-based doughs were correlated. We extracted total RNA from both a liquid and a flour-based dough. We used Northern blotting to analyze mRNA levels of three stress marker genes, HSP26, GPD1, and ENA1, and 10 genes in different metabolic subcategories. All 13 genes had the same transcriptional profile in both systems. Hence, the model appears to effectively mimic the environment encountered by baker's yeast in high-sugar dough. The liquid dough can be used to help understand the connections between technological traits and biological functions and to facilitate studies of gene expression under commercially important, but experimentally intractable, conditions.Comisión Interministerial de Ciencia y Tecnología project (AGL2001-1203

miRNomic Signature in Very Low Birth-Weight Neonates Discriminates Late-Onset Gram-Positive Sepsis from Controls

Background and Objectives. Neonatal sepsis is a serious condition with a high rate of mortality and morbidity. Currently, the gold standard for sepsis diagnosis is a positive blood culture, which takes 48–72 h to yield results. We hypothesized that identifying differentially expressed miRNA pattern in neonates with late-onset Gram-positive sepsis would help with an earlier diagnosis and therapy. Methods. This is a prospective observational study in newborn infants with late-onset Gram positive bacterial sepsis and non-septic controls. Complementary to blood culture, an aliquot of 0.5 mL of blood was used to determine small non-coding RNA expression profiling using the GeneChip miRNA 4.0 Array. Results. A total of 11 very low birth-weight neonates with late-onset Gram-positive sepsis and 16 controls were analyzed. Further, 217 differentially expressed miRNAs were obtained between both groups. Subsequently, a combined analysis was performed with these miRNAs and 4297 differentially expressed genes. We identified 33 miRNAs that regulate our mRNAs, and the most relevant biological processes are associated with the immune system and the inflammatory response. Conclusions. The miRNA profiling in very low birth-weight neonates distinguishes late-onset Gram-positive sepsis versus control neonates

Método para mejorar la viabilidad y la capacidad fermentativa de levaduras de panadería en condiciones de estrés hiperosmótico y estrés por congelación

Fecha de presentación internacional: 03.08.2004. - Titular: Consejo Superior de Investigaciones Científicas (CSIC)[EN] The invention relates to a method of improving the viability and the fermentation capacity of baking yeasts under hyperosmotic stress and freezing stress conditions, through the genetic transformation of said yeasts using a nucleotide sequence that enables the expression of a protein with Crzl activity.[ES] La presente invención se refiere a un método para mejorar la viabilidad y la capacidad fermentativa de levaduras de panadería en condiciones de estrés hiperosmótico y estrés por congelación, mediante la transformación genética de las levaduras con una secuencia de nucleótidos que permite la expresión de una proteína con actividad Crzl.Peer reviewe

A downshift in temperature activates the high osmolarity glycerol (HOG) pathway, which determines freeze tolerance in Saccharomyces cerevisiae

The molecular mechanisms that enable yeast cells to detect and transmit cold signals and their physiological significance in the adaptive response to low temperatures are unknown. Here, we have demonstrated that the MAPK Hog1p is specifically activated in response to cold. Phosphorylation of Hog1p was dependent on Pbs2p, the MAPK kinase (MAPKK) of the high osmolarity glycerol (HOG) pathway, and Ssk1p, the response regulator of the two-component system Sln1p-Ypd1p. However, Sho1p was not required. Interestingly, phosphorylation of Hog1p was stimulated at 30°C in cells exposed to the membrane rigidifier agent dimethyl sulfoxide. Moreover, Hog1p activation occurred specifically through the Sln1 branch. This suggests that Sln1p monitors changes in membrane fluidity caused by cold. Quite remarkably, activation of Hog1p at low temperatures affected the transcriptional response to cold shock. Indeed, the absence of Hog1p impaired the cold-instigated expression of genes for trehalose- and glycerol-synthesizing enzymes and small chaperones. Moreover, a downward transfer to 12 or 4°C stimulated the overproduction of glycerol in a Hog1p-dependent manner. However, hog1Δ mutant cells showed no growth defects at 12°C as compared with the wild type. On the contrary, deletion of HOG1 or GPD1 decreased tolerance to freezing of wildtype cells preincubated at a low temperature, whereas no differences could be detected in cells shifted directly from 30 to -20°C. Thus, exposure to low temperatures triggered a Hog1p-dependent accumulation of glycerol, which is essential for freeze protection. © 2006 by The American Society for Biochemistry and Molecular Biology, Inc.Peer Reviewe

Overexpression of the Calcineurin Target CRZ1 Provides Freeze Tolerance and Enhances the Fermentative Capacity of Baker's Yeast▿

Recent years have shown a huge growth in the market of industrial baker's yeasts (Saccharomyces cerevisiae), with the need for strains affording better performance in prefrozen dough. Evidence suggests that during the freezing process, cells can suffer biochemical damage caused by osmotic stress. Nevertheless, the involvement of ion-responsive transcriptional factors and pathways in conferring freeze resistance has not yet been examined. Here, we have investigated the role of the salt-responsive calcineurin-Crz1p pathway in mediating tolerance to freezing by industrial baker's yeast. Overexpression of CRZ1 in the industrial HS13 strain increased both salt and freeze tolerance and improved the leavening ability of baker's yeast in high-sugar dough. Moreover, engineered cells were able to produce more gas during fermentation of prefrozen dough than the parental strain. Similar effects were observed for overexpression of TdCRZ1, the homologue to CRZ1 in Torulaspora delbrueckii, suggesting that expression of calcineurin-Crz1p target genes can alleviate the harmful effects of ionic stress during freezing. However, overexpression of STZ and FTZ, two unrelated Arabidopsis thaliana genes encoding Cys2/His2-type zinc finger proteins, also conferred freeze resistance in yeast. Furthermore, experiments with Δcnb1 and Δcrz1 mutants failed to show a freeze-sensitive phenotype, even in cells pretreated with NaCl. Overall, our results demonstrate that overexpression of CRZ1 has the potential to be a useful tool for increasing freeze tolerance and fermentative capacity in industrial strains. However, these effects do not appear to be mediated through activation of known salt-responding pathways