Luciferase NanoLuc as a reporter for gene expression and protein levels in Saccharomyces cerevisiae.

PubMed

Masser, Anna E; Kandasamy, Ganapathi; Kaimal, Jayasankar Mohanakrishnan; Andréasson, Claes

2016-05-01

Reporter proteins are essential tools in the study of biological processes and are employed to monitor changes in gene expression and protein levels. Luciferases are reporter proteins that enable rapid and highly sensitive detection with an outstanding dynamic range. Here we evaluated the usefulness of the 19 kDa luciferase NanoLuc (Nluc), derived from the deep sea shrimp Oplophorus gracilirostris, as a reporter protein in yeast. Cassettes with codon-optimized genes expressing yeast Nluc (yNluc) or its destabilized derivative yNlucPEST have been assembled in the context of the dominant drug resistance marker kanMX. The reporter proteins do not impair the growth of yeast cells and exhibit half-lives of 40 and 5 min, respectively. The commercial substrate Nano-Glo® is compatible with detection of yNluc bioluminescence in < 50 cells. Using the unstable yNlucPEST to report on the rapid and transient expression of a heat-shock promoter (PCYC1-HSE ), we found a close match between the intensity of the bioluminescent signal and mRNA levels during both induction and decay. We demonstrated that the bioluminescence of yNluc fused to the C-terminus of a temperature-sensitive protein reports on its protein levels. In conclusion, yNluc and yNlucPEST are valuable new reporter proteins suitable for experiments with yeast using standard commercial substrate. © 2016 The Authors. Yeast published by John Wiley & Sons Ltd. © 2016 The Authors. Yeast published by John Wiley & Sons Ltd.

Macromitophagy is a longevity assurance process that in chronologically aging yeast limited in calorie supply sustains functional mitochondria and maintains cellular lipid homeostasis

PubMed Central

Burstein, Michelle T.; Koupaki, Olivia; Gomez-Perez, Alejandra; Levy, Sean; Pluska, Lukas; Mattie, Sevan; Rafeh, Rami; Iouk, Tatiana; Sheibani, Sara; Greenwood, Michael; Vali, Hojatollah; Titorenko, Vladimir I.

2013-01-01

Macromitophagy controls mitochondrial quality and quantity. It involves the sequestration of dysfunctional or excessive mitochondria within double-membrane autophagosomes, which then fuse with the vacuole/lysosome to deliver these mitochondria for degradation. To investigate a physiological role of macromitophagy in yeast, we examined how the atg32Δ-dependent mutational block of this process influences the chronological lifespan of cells grown in a nutrient-rich medium containing low (0.2%) concentration of glucose. Under these longevity-extending conditions of caloric restriction (CR) yeast cells are not starving. We also assessed a role of macromitophagy in lifespan extension by lithocholic acid (LCA), a bile acid that prolongs yeast longevity under CR conditions. Our findings imply that macromitophagy is a longevity assurance process underlying the synergistic beneficial effects of CR and LCA on yeast lifespan. Our analysis of how the atg32Δ mutation influences mitochondrial morphology, composition and function revealed that macromitophagy is required to maintain a network of healthy mitochondria. Our comparative analysis of the membrane lipidomes of organelles purified from wild-type and atg32Δ cells revealed that macromitophagy is required for maintaining cellular lipid homeostasis. We concluded that macromitophagy defines yeast longevity by modulating vital cellular processes inside and outside of mitochondria. PMID:23553280

Gateway Vectors for Efficient Artificial Gene Assembly In Vitro and Expression in Yeast Saccharomyces cerevisiae

PubMed Central

Giuraniuc, Claudiu V.; MacPherson, Murray; Saka, Yasushi

2013-01-01

Construction of synthetic genetic networks requires the assembly of DNA fragments encoding functional biological parts in a defined order. Yet this may become a time-consuming procedure. To address this technical bottleneck, we have created a series of Gateway shuttle vectors and an integration vector, which facilitate the assembly of artificial genes and their expression in the budding yeast Saccharomyces cerevisiae. Our method enables the rapid construction of an artificial gene from a promoter and an open reading frame (ORF) cassette by one-step recombination reaction in vitro. Furthermore, the plasmid thus created can readily be introduced into yeast cells to test the assembled gene’s functionality. As flexible regulatory components of a synthetic genetic network, we also created new versions of the tetracycline-regulated transactivators tTA and rtTA by fusing them to the auxin-inducible degron (AID). Using our gene assembly approach, we made yeast expression vectors of these engineered transactivators, AIDtTA and AIDrtTA and then tested their functions in yeast. We showed that these factors can be regulated by doxycycline and degraded rapidly after addition of auxin to the medium. Taken together, the method for combinatorial gene assembly described here is versatile and would be a valuable tool for yeast synthetic biology. PMID:23675537

Plant-Derived Transcription Factors for Orthologous Regulation of Gene Expression in the Yeast Saccharomyces cerevisiae.

PubMed

Naseri, Gita; Balazadeh, Salma; Machens, Fabian; Kamranfar, Iman; Messerschmidt, Katrin; Mueller-Roeber, Bernd

2017-09-15

Control of gene expression by transcription factors (TFs) is central in many synthetic biology projects for which a tailored expression of one or multiple genes is often needed. As TFs from evolutionary distant organisms are unlikely to affect gene expression in a host of choice, they represent excellent candidates for establishing orthogonal control systems. To establish orthogonal regulators for use in yeast (Saccharomyces cerevisiae), we chose TFs from the plant Arabidopsis thaliana. We established a library of 106 different combinations of chromosomally integrated TFs, activation domains (yeast GAL4 AD, herpes simplex virus VP64, and plant EDLL) and synthetic promoters harboring cognate cis-regulatory motifs driving a yEGFP reporter. Transcriptional output of the different driver/reporter combinations varied over a wide spectrum, with EDLL being a considerably stronger transcription activation domain in yeast than the GAL4 activation domain, in particular when fused to Arabidopsis NAC TFs. Notably, the strength of several NAC-EDLL fusions exceeded that of the strong yeast TDH3 promoter by 6- to 10-fold. We furthermore show that plant TFs can be used to build regulatory systems encoded by centromeric or episomal plasmids. Our library of TF-DNA binding site combinations offers an excellent tool for diverse synthetic biology applications in yeast.

Mammalian amyloidogenic proteins promote prion nucleation in yeast.

PubMed

Chandramowlishwaran, Pavithra; Sun, Meng; Casey, Kristin L; Romanyuk, Andrey V; Grizel, Anastasiya V; Sopova, Julia V; Rubel, Aleksandr A; Nussbaum-Krammer, Carmen; Vorberg, Ina M; Chernoff, Yury O

2018-03-02

Fibrous cross-β aggregates (amyloids) and their transmissible forms (prions) cause diseases in mammals (including humans) and control heritable traits in yeast. Initial nucleation of a yeast prion by transiently overproduced prion-forming protein or its (typically, QN-rich) prion domain is efficient only in the presence of another aggregated (in most cases, QN-rich) protein. Here, we demonstrate that a fusion of the prion domain of yeast protein Sup35 to some non-QN-rich mammalian proteins, associated with amyloid diseases, promotes nucleation of Sup35 prions in the absence of pre-existing aggregates. In contrast, both a fusion of the Sup35 prion domain to a multimeric non-amyloidogenic protein and the expression of a mammalian amyloidogenic protein that is not fused to the Sup35 prion domain failed to promote prion nucleation, further indicating that physical linkage of a mammalian amyloidogenic protein to the prion domain of a yeast protein is required for the nucleation of a yeast prion. Biochemical and cytological approaches confirmed the nucleation of protein aggregates in the yeast cell. Sequence alterations antagonizing or enhancing amyloidogenicity of human amyloid-β (associated with Alzheimer's disease) and mouse prion protein (associated with prion diseases), respectively, antagonized or enhanced nucleation of a yeast prion by these proteins. The yeast-based prion nucleation assay, developed in our work, can be employed for mutational dissection of amyloidogenic proteins. We anticipate that it will aid in the identification of chemicals that influence initial amyloid nucleation and in searching for new amyloidogenic proteins in a variety of proteomes. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Autolytic activity of human calpain 7 is enhanced by ESCRT-III-related protein IST1 through MIT-MIM interaction.

PubMed

Osako, Yohei; Maemoto, Yuki; Tanaka, Ryohei; Suzuki, Hironori; Shibata, Hideki; Maki, Masatoshi

2010-11-01

Calpain 7, a mammalian ortholog of yeast Cpl1/Rim13 and fungal PalB, is an atypical calpain that lacks a penta-EF-hand domain. Previously, we reported that a region containing a tandem repeat of microtubule-interacting and transport (MIT) domains in calpain 7 interacts with a subset of endosomal sorting complex required for transport (ESCRT)-III-related proteins, suggesting involvement of calpain 7 in the ESCRT system. Although yeast and fungal calpains are thought to be involved in alkaline adaptation via limited proteolysis of specific transcription factors, proteolytic activity of calpain 7 has not been demonstrated yet. In this study, we investigated the interaction between calpain 7 and a newly reported ESCRT-III family member, increased sodium tolerance-1 (IST1), which possesses two different types of MIT-interacting motifs (MIM1 and MIM2). We found that glutathione-S-transferase (GST)-fused tandem MIT domains of calpain 7 (calpain 7MIT) pulled down FLAG-tagged IST1 expressed in HEK293T cells. Coimmunoprecipitation assays with various deletion or point mutants of epitope-tagged calpain 7 and IST1 revealed that both repetitive MIT domains and MIMs are required for efficient interaction. Direct MIT-MIM binding was confirmed by a pulldown experiment with GST-fused IST1 MIM and purified recombinant calpain 7MIT. Furthermore, we found that the GST-MIM protein enhances the autolysis of purified Strep-tagged monomeric green fluorescent protein (mGFP)-fused calpain 7 (mGFP-calpain 7-Strep). The autolysis was almost completely abolished by 10 mmN-ethylmaleimide but only partially inhibited by 1 mm leupeptin or E-64. The putative catalytic Cys290-substituted mutant (mGFP-calpain 7(C290S)-Strep) showed no autolytic activity. These results demonstrate for the first time that human calpain 7 is proteolytically active, and imply that calpain 7 is activated in the ESCRT system. © 2010 The Authors Journal compilation © 2010 FEBS.

Tracking Diacylglycerol and Phosphatidic Acid Pools in Budding Yeast

PubMed Central

Ganesan, Suriakarthiga; Shabits, Brittney N.; Zaremberg, Vanina

2015-01-01

Phosphatidic acid (PA) and diacylglycerol (DAG) are key signaling molecules and important precursors for the biosynthesis of all glycerolipids found in eukaryotes. Research conducted in the model organism Saccharomyces cerevisiae has been at the forefront of the identification of the enzymes involved in the metabolism and transport of PA and DAG. Both these lipids can alter the local physical properties of membranes by introducing negative curvature, but the anionic nature of the phosphomonoester headgroup in PA sets it apart from DAG. As a result, the mechanisms underlying PA and DAG interaction with other lipids and proteins are notoriously different. This is apparent from the analysis of the protein domains responsible for recognition and binding to each of these lipids. We review the current evidence obtained using the PA-binding proteins and domains fused to fluorescent proteins for in vivo tracking of PA pools in yeast. In addition, we present original results for visualization of DAG pools in yeast using the C1 domain from mammalian PKCδ. An emerging first cellular map of the distribution of PA and DAG pools in actively growing yeast is discussed. PMID:27081314

Convergent evolution of a fused sexual cycle promotes the haploid lifestyle

NASA Astrophysics Data System (ADS)

Sherwood, Racquel Kim; Scaduto, Christine M.; Torres, Sandra E.; Bennett, Richard J.

2014-02-01

Sexual reproduction is restricted to eukaryotic species and involves the fusion of haploid gametes to form a diploid cell that subsequently undergoes meiosis to generate recombinant haploid forms. This process has been extensively studied in the unicellular yeast Saccharomyces cerevisiae, which exhibits separate regulatory control over mating and meiosis. Here we address the mechanism of sexual reproduction in the related hemiascomycete species Candida lusitaniae. We demonstrate that, in contrast to S. cerevisiae, C. lusitaniae exhibits a highly integrated sexual program in which the programs regulating mating and meiosis have fused. Profiling of the C. lusitaniae sexual cycle revealed that gene expression patterns during mating and meiosis were overlapping, indicative of co-regulation. This was particularly evident for genes involved in pheromone MAPK signalling, which were highly induced throughout the sexual cycle of C. lusitaniae. Furthermore, genetic analysis showed that the orthologue of IME2, a `diploid-specific' factor in S. cerevisiae, and STE12, the master regulator of S. cerevisiae mating, were each required for progression through both mating and meiosis in C. lusitaniae. Together, our results establish that sexual reproduction has undergone significant rewiring between S. cerevisiae and C. lusitaniae, and that a concerted sexual cycle operates in C. lusitaniae that is more reminiscent of the distantly related ascomycete, Schizosaccharomyces pombe. We discuss these results in light of the evolution of sexual reproduction in yeast, and propose that regulatory coupling of mating and meiosis has evolved multiple times as an adaptation to promote the haploid lifestyle.

Improved bioethanol production using fusants of Saccharomyces cerevisiae and xylose-fermenting yeasts.

PubMed

Kumari, Rajni; Pramanik, K

2012-06-01

The present research deals with the development of a hybrid yeast strain with the aim of converting pentose and hexose sugar components of lignocellulosic substrate to bioethanol by fermentation. Different fusant strains were obtained by fusing protoplasts of Saccharomyces cerevisiae and xylose-fermenting yeasts such as Pachysolen tannophilus, Candida shehatae and Pichia stipitis. The fusants were sorted by fluorescent-activated cell sorter and further confirmed by molecular characterization. The fusants were evaluated by fermentation of glucose-xylose mixture and the highest ethanol producing fusant was used for further study to ferment hydrolysates produced by acid pretreatment and enzymatic hydrolysis of cotton gin waste. Among the various fusant and parental strains used under present study, RPR39 was found to be stable and most efficient strain giving maximum ethanol concentration (76.8 ± 0.31 g L(-1)), ethanol productivity (1.06 g L(-1) h(-1)) and ethanol yield (0.458 g g(-1)) by fermentation of glucose-xylose mixture under test conditions. The fusant has also shown encouraging result in fermenting hydrolysates of cotton gin waste with ethanol concentration of 7.08 ± 0.142 g L(-1), ethanol yield of 0.44 g g(-1), productivity of 0.45 g L(-1) h(-1) and biomass yield of 0.40 g g(-1).

Fusion, fission, and transport control asymmetric inheritance of mitochondria and protein aggregates

PubMed Central

Böckler, Stefan; Chelius, Xenia; Hock, Nadine; Weiss, Matthias

2017-01-01

Partitioning of cell organelles and cytoplasmic components determines the fate of daughter cells upon asymmetric division. We studied the role of mitochondria in this process using budding yeast as a model. Anterograde mitochondrial transport is mediated by the myosin motor, Myo2. A genetic screen revealed an unexpected interaction of MYO2 and genes required for mitochondrial fusion. Genetic analyses, live-cell microscopy, and simulations in silico showed that fused mitochondria become critical for inheritance and transport across the bud neck in myo2 mutants. Similarly, fused mitochondria are essential for retention in the mother when bud-directed transport is enforced. Inheritance of a less than critical mitochondrial quantity causes a severe decline of replicative life span of daughter cells. Myo2-dependent mitochondrial distribution also is critical for the capture of heat stress–induced cytosolic protein aggregates and their retention in the mother cell. Together, these data suggest that coordination of mitochondrial transport, fusion, and fission is critical for asymmetric division and rejuvenation of daughter cells. PMID:28615194

Synthetic Core Promoters as Universal Parts for Fine-Tuning Expression in Different Yeast Species

PubMed Central

2016-01-01

Synthetic biology and metabolic engineering experiments frequently require the fine-tuning of gene expression to balance and optimize protein levels of regulators or metabolic enzymes. A key concept of synthetic biology is the development of modular parts that can be used in different contexts. Here, we have applied a computational multifactor design approach to generate de novo synthetic core promoters and 5′ untranslated regions (UTRs) for yeast cells. In contrast to upstream cis-regulatory modules (CRMs), core promoters are typically not subject to specific regulation, making them ideal engineering targets for gene expression fine-tuning. 112 synthetic core promoter sequences were designed on the basis of the sequence/function relationship of natural core promoters, nucleosome occupancy and the presence of short motifs. The synthetic core promoters were fused to the Pichia pastoris AOX1 CRM, and the resulting activity spanned more than a 200-fold range (0.3% to 70.6% of the wild type AOX1 level). The top-ten synthetic core promoters with highest activity were fused to six additional CRMs (three in P. pastoris and three in Saccharomyces cerevisiae). Inducible CRM constructs showed significantly higher activity than constitutive CRMs, reaching up to 176% of natural core promoters. Comparing the activity of the same synthetic core promoters fused to different CRMs revealed high correlations only for CRMs within the same organism. These data suggest that modularity is maintained to some extent but only within the same organism. Due to the conserved role of eukaryotic core promoters, this rational design concept may be transferred to other organisms as a generic engineering tool. PMID:27973777

ER-associated SNAREs and Sey1p mediate nuclear fusion at two distinct steps during yeast mating.

PubMed

Rogers, Jason V; Arlow, Tim; Inkellis, Elizabeth R; Koo, Timothy S; Rose, Mark D

2013-12-01

During yeast mating, two haploid nuclei fuse membranes to form a single diploid nucleus. However, the known proteins required for nuclear fusion are unlikely to function as direct fusogens (i.e., they are unlikely to directly catalyze lipid bilayer fusion) based on their predicted structure and localization. Therefore we screened known fusogens from vesicle trafficking (soluble N-ethylmaleimide-sensitive factor attachment protein receptors [SNAREs]) and homotypic endoplasmic reticulum (ER) fusion (Sey1p) for additional roles in nuclear fusion. Here we demonstrate that the ER-localized SNAREs Sec20p, Ufe1p, Use1p, and Bos1p are required for efficient nuclear fusion. In contrast, Sey1p is required indirectly for nuclear fusion; sey1Δ zygotes accumulate ER at the zone of cell fusion, causing a block in nuclear congression. However, double mutants of Sey1p and Sec20p, Ufe1p, or Use1p, but not Bos1p, display extreme ER morphology defects, worse than either single mutant, suggesting that retrograde SNAREs fuse ER in the absence of Sey1p. Together these data demonstrate that SNAREs mediate nuclear fusion, ER fusion after cell fusion is necessary to complete nuclear congression, and there exists a SNARE-mediated, Sey1p-independent ER fusion pathway.

Integrated Approach To Producing High-Purity Trehalose from Maltose by the Yeast Yarrowia lipolytica Displaying Trehalose Synthase (TreS) on the Cell Surface.

PubMed

Li, Ning; Wang, Hengwei; Li, Lijuan; Cheng, Huiling; Liu, Dawen; Cheng, Hairong; Deng, Zixin

2016-08-10

An alternative strategy that integrated enzyme production, trehalose biotransformation, and bioremoval in one bioreactor was developed in this study, thus simplifying the traditional procedures used for trehalose production. The trehalose synthase gene from a thermophilic archaea, Picrophilus torridus, was first fused to the YlPir1 anchor gene and then inserted into the genome of Yarrowia lipolytica, thus yielding an engineered yeast strain. The trehalose yield reached 73% under optimal conditions. The thermal and pH stabilities of the displayed enzyme were improved compared to those of its free form purified from recombinant Escherichia coli. After biotransformation, the glucose byproduct and residual maltose were directly fermented to ethanol by a Saccharomyces cerevisiae strain. Ethanol can be separated by distillation, and high-purity trehalose can easily be obtained from the fermentation broth. The results show that this one-pot procedure is an efficient approach to the economical production of trehalose from maltose.

Synthesis of a beta-estradiol-biotin chimera that potently heterodimerizes estrogen receptor and streptavidin proteins in a yeast three-hybrid system.

PubMed

Hussey, Stephen L; Muddana, Smita S; Peterson, Blake R

2003-04-02

Small molecules that dimerize proteins in living cells provide powerful probes of biological processes and have potential as tools for the identification of protein targets of natural products. We synthesized 7-alpha-substituted derivatives of beta-estradiol tethered to the natural product biotin to regulate heterodimerization of estrogen receptor (ER) and streptavidin (SA) proteins expressed as components of a yeast three-hybrid system. Addition of an estradiol-biotin chimera bearing a 19-atom linker to yeast expressing DNA-bound ER-alpha or ER-beta LexA fusion proteins and wild-type SA protein fused to the B42 activation domain activated reporter gene expression by as much as 450-fold in vivo (10 muM ligand). Comparative analysis of lower affinity Y43A (biotin Kd approximately 100 pM) and W120A (biotin Kd approximately 100 nM) mutants of SA indicated that moderate affinity interactions can be readily detected with this system. Comparison of a 7-alpha-substituted estradiol-biotin chimera with a structurally similar dexamethasone-biotin chimera revealed that yeast expressing ER proteins can detect cognate ligands with up to 5-fold greater potency and 70-fold higher activity than yeast expressing analogous glucocorticoid receptor (GR) proteins. This approach may facilitate the identification of protein targets of biologically active small molecules screened against genetically encoded libraries of proteins expressed in yeast three-hybrid systems.

Anchoring plant metallothioneins to the inner face of the plasma membrane of Saccharomyces cerevisiae cells leads to heavy metal accumulation.

PubMed

Ruta, Lavinia Liliana; Lin, Ya-Fen; Kissen, Ralph; Nicolau, Ioana; Neagoe, Aurora Daniela; Ghenea, Simona; Bones, Atle M; Farcasanu, Ileana Cornelia

2017-01-01

In this study we engineered yeast cells armed for heavy metal accumulation by targeting plant metallothioneins to the inner face of the yeast plasma membrane. Metallothioneins (MTs) are cysteine-rich proteins involved in the buffering of excess metal ions, especially Cu(I), Zn(II) or Cd(II). The cDNAs of seven Arabidopsis thaliana MTs (AtMT1a, AtMT1c, AtMT2a, AtMT2b, AtMT3, AtMT4a and AtMT4b) and four Noccaea caerulescens MTs (NcMT1, NcMT2a, NcMT2b and NcMT3) were each translationally fused to the C-terminus of a myristoylation green fluorescent protein variant (myrGFP) and expressed in Saccharomyces cerevisiae cells. The myrGFP cassette introduced a yeast myristoylation sequence which allowed directional targeting to the cytosolic face of the plasma membrane along with direct monitoring of the intracellular localization of the recombinant protein by fluorescence microscopy. The yeast strains expressing plant MTs were investigated against an array of heavy metals in order to identify strains which exhibit the (hyper)accumulation phenotype without developing toxicity symptoms. Among the transgenic strains which could accumulate Cu(II), Zn(II) or Cd(II), but also non-canonical metal ions, such as Co(II), Mn(II) or Ni(II), myrGFP-NcMT3 qualified as the best candidate for bioremediation applications, thanks to the robust growth accompanied by significant accumulative capacity.

Roles of the checkpoint sensor clamp Rad9-Rad1-Hus1 (911)-complex and the clamp loaders Rad17-RFC and Ctf18-RFC in Schizosaccharomyces pombe telomere maintenance.

PubMed

Khair, Lyne; Chang, Ya-Ting; Subramanian, Lakxmi; Russell, Paul; Nakamura, Toru M

2010-06-01

While telomeres must provide mechanisms to prevent DNA repair and DNA damage checkpoint factors from fusing chromosome ends and causing permanent cell cycle arrest, these factors associate with functional telomeres and play critical roles in the maintenance of telomeres. Previous studies have established that Tel1 (ATM) and Rad3 (ATR) kinases play redundant but essential roles for telomere maintenance in fission yeast. In addition, the Rad9-Rad1-Hus1 (911) and Rad17-RFC complexes work downstream of Rad3 (ATR) in fission yeast telomere maintenance. Here, we investigated how 911, Rad17-RFC and another RFC-like complex Ctf18-RFC contribute to telomere maintenance in fission yeast cells lacking Tel1 and carrying a novel hypomorphic allele of rad3 (DBD-rad3), generated by the fusion between the DNA binding domain (DBD) of the fission yeast telomere capping protein Pot1 and Rad3. Our investigations have uncovered a surprising redundancy for Rad9 and Hus1 in allowing Rad1 to contribute to telomere maintenance in DBD-rad3 tel1 cells. In addition, we found that Rad17-RFC and Ctf18-RFC carry out redundant telomere maintenance functions in DBD-rad3 tel1 cells. Since checkpoint sensor proteins are highly conserved, genetic redundancies uncovered here may be relevant to telomere maintenance and detection of DNA damage in other eukaryotes.

Roles of the Checkpoint Sensor Clamp Rad9-Rad1-Hus1 (911)-Complex and the Clamp Loaders Rad17-RFC and Ctf18-RFC in Schizosaccharomyces pombe Telomere Maintenance

PubMed Central

Khair, Lyne; Chang, Ya-Ting; Subramanian, Lakxmi; Russell, Paul; Nakamura, Toru M.

2011-01-01

While telomeres must provide mechanisms to prevent DNA repair and DNA damage checkpoint factors from fusing chromosome ends and causing permanent cell cycle arrest, these factors associate with functional telomeres and play critical roles in the maintenance of telomeres. Previous studies have established that Tel1 (ATM) and Rad3 (ATR) kinases play redundant but essential roles for telomere maintenance in fission yeast. In addition, the Rad9-Rad1-Hus1 (911) and Rad17-RFC complexes work downstream of Rad3 (ATR) in fission yeast telomere maintenance. Here, we investigated how 911, Rad17-RFC and another RFC-like complex Ctf18-RFC contribute to telomere maintenance in fission yeast cells lacking Tel1 and carrying a novel hypomorphic allele of rad3 (DBD-rad3), generated by the fusion between the DNA binding domain (DBD) of the fission yeast telomere capping protein Pot1 and Rad3. Our investigations have uncovered a surprising redundancy for Rad9 and Hus1 in allowing Rad1 to contribute to telomere maintenance in DBD-rad3 tel1Δ cells. In addition, we found that Rad17-RFC and Ctf18-RFC carry out redundant telomere maintenance functions in DBD-rad3 tel1Δ cells. Since checkpoint sensor proteins are highly conserved, genetic redundancies uncovered here may be relevant to telomere maintenance and detection of DNA damage in other eukaryotes. PMID:20505337

Survey of molecular chaperone requirement for the biosynthesis of hamster polyomavirus VP1 protein in Saccharomyces cerevisiae.

PubMed

Valaviciute, Monika; Norkiene, Milda; Goda, Karolis; Slibinskas, Rimantas; Gedvilaite, Alma

2016-07-01

A number of viruses utilize molecular chaperones during various stages of their life cycle. It has been shown that members of the heat-shock protein 70 (Hsp70) chaperone family assist polyomavirus capsids during infection. However, the molecular chaperones that assist the formation of recombinant capsid viral protein 1 (VP1)-derived virus-like particles (VLPs) in yeast remain unclear. A panel of yeast strains with single chaperone gene deletions were used to evaluate the chaperones required for biosynthesis of recombinant hamster polyomavirus capsid protein VP1. The impact of deletion or mild overexpression of chaperone genes was determined in live cells by flow cytometry using enhanced green fluorescent protein (EGFP) fused with VP1. Targeted genetic analysis demonstrated that VP1-EGFP fusion protein levels were significantly higher in yeast strains in which the SSZ1 or ZUO1 genes encoding ribosome-associated complex components were deleted. The results confirmed the participation of cytosolic Hsp70 chaperones and suggested the potential involvement of the Ydj1 and Caj1 co-chaperones and the endoplasmic reticulum chaperones in the biosynthesis of VP1 VLPs in yeast. Likewise, the markedly reduced levels of VP1-EGFP in Δhsc82 and Δhsp82 yeast strains indicated that both Hsp70 and Hsp90 chaperones might assist VP1 VLPs during protein biosynthesis.

Use of the Yeast Pichia pastoris as an Expression Host for Secretion of Enterocin L50, a Leaderless Two-Peptide (L50A and L50B) Bacteriocin from Enterococcus faecium L50▿

PubMed Central

Basanta, Antonio; Gómez-Sala, Beatriz; Sánchez, Jorge; Diep, Dzung B.; Herranz, Carmen; Hernández, Pablo E.; Cintas, Luis M.

2010-01-01

In this work, we report the expression and secretion of the leaderless two-peptide (EntL50A and EntL50B) bacteriocin enterocin L50 from Enterococcus faecium L50 by the methylotrophic yeast Pichia pastoris X-33. The bacteriocin structural genes entL50A and entL50B were fused to the Saccharomyces cerevisiae gene region encoding the mating pheromone α-factor 1 secretion signal (MFα1s) and cloned, separately and together (entL50AB), into the P. pastoris expression and secretion vector pPICZαA, which contains the methanol-inducible alcohol oxidase promoter (PAOX1) to express the fusion genes. After transfer into the yeast, the recombinant plasmids were integrated into the genome, resulting in three bacteriocinogenic yeast strains able to produce and secrete the individual bacteriocin peptides EntL50A and EntL50B separately and together. The secretion was efficiently directed by MFα1s through the Sec system, and the precursor peptides were found to be correctly processed to form mature and active bacteriocin peptides. The present work describes for the first time the heterologous expression and secretion of a two-peptide non-pediocin-like bacteriocin by a yeast. PMID:20348300

Yeast cell surface display for lipase whole cell catalyst and its applications

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Yun; Zhang, Rui; Lian, Zhongshuai

The cell surface display technique allows for the expression of target proteins or peptides on the microbial cell surface by fusing an appropriate protein as an anchoring motif. Yeast display systems, such as Pichia pastoris, Yarowia lipolytica and Saccharomyces cerevisiae, are ideal, alternative and extensive display systems with the advantage of simple genetic manipulation and post-translational modification of expressed heterologous proteins. Engineered yeasts show high performance characteristics and variant utilizations. Herein, we comprehensively summarize the variant factors affecting lipase whole cell catalyst activity and display efficiency, including the structure and size of target proteins, screening anchor proteins, type and chainmore » length of linkers, and the appropriate matching rules among the above-mentioned display units. Furthermore, we also address novel approaches to enhance stability and activity of recombinant lipases, such as VHb gene co-expression, multi-enzyme co-display technique, and the micro-environmental interference and self-assembly techniques. Finally, we represent the variety of applications of whole cell surface displayed lipases on yeast cells in non-aqueous phases, including synthesis of esters, PUFA enrichment, resolution of chiral drugs, organic synthesis and biofuels. We demonstrate that the lipase surface display technique is a powerful tool for functionalizing yeasts to serve as whole cell catalysts, and increasing interest is providing an impetus for broad application of this technique.« less

ER-associated SNAREs and Sey1p mediate nuclear fusion at two distinct steps during yeast mating

PubMed Central

Rogers, Jason V.; Arlow, Tim; Inkellis, Elizabeth R.; Koo, Timothy S.; Rose, Mark D.

2013-01-01

During yeast mating, two haploid nuclei fuse membranes to form a single diploid nucleus. However, the known proteins required for nuclear fusion are unlikely to function as direct fusogens (i.e., they are unlikely to directly catalyze lipid bilayer fusion) based on their predicted structure and localization. Therefore we screened known fusogens from vesicle trafficking (soluble N-ethylmaleimide–sensitive factor attachment protein receptors [SNAREs]) and homotypic endoplasmic reticulum (ER) fusion (Sey1p) for additional roles in nuclear fusion. Here we demonstrate that the ER-localized SNAREs Sec20p, Ufe1p, Use1p, and Bos1p are required for efficient nuclear fusion. In contrast, Sey1p is required indirectly for nuclear fusion; sey1Δ zygotes accumulate ER at the zone of cell fusion, causing a block in nuclear congression. However, double mutants of Sey1p and Sec20p, Ufe1p, or Use1p, but not Bos1p, display extreme ER morphology defects, worse than either single mutant, suggesting that retrograde SNAREs fuse ER in the absence of Sey1p. Together these data demonstrate that SNAREs mediate nuclear fusion, ER fusion after cell fusion is necessary to complete nuclear congression, and there exists a SNARE-mediated, Sey1p-independent ER fusion pathway. PMID:24152736

Maltose-Binding Protein Enhances Secretion of Recombinant Human Granzyme B Accompanied by In Vivo Processing of a Precursor MBP Fusion Protein

PubMed Central

Dälken, Benjamin; Jabulowsky, Robert A.; Oberoi, Pranav; Benhar, Itai; Wels, Winfried S.

2010-01-01

Background The apoptosis-inducing serine protease granzyme B (GrB) is an important factor contributing to lysis of target cells by cytotoxic lymphocytes. Expression of enzymatically active GrB in recombinant form is a prerequisite for functional analysis and application of GrB for therapeutic purposes. Methods and Findings We investigated the influence of bacterial maltose-binding protein (MBP) fused to GrB via a synthetic furin recognition motif on the expression of the MBP fusion protein also containing an N-terminal α-factor signal peptide in the yeast Pichia pastoris. MBP markedly enhanced the amount of GrB secreted into culture supernatant, which was not the case when GrB was fused to GST. MBP-GrB fusion protein was cleaved during secretion by an endogenous furin-like proteolytic activity in vivo, liberating enzymatically active GrB without the need of subsequent in vitro processing. Similar results were obtained upon expression of a recombinant fragment of the ErbB2/HER2 receptor protein or GST as MBP fusions. Conclusions Our results demonstrate that combination of MBP as a solubility enhancer with specific in vivo cleavage augments secretion of processed and functionally active proteins from yeast. This strategy may be generally applicable to improve folding and increase yields of recombinant proteins. PMID:21203542

Pet10p is a yeast perilipin that stabilizes lipid droplets and promotes their assembly

PubMed Central

Kinch, Lisa N.; Grishin, Nick V.

2017-01-01

Pet10p is a yeast lipid droplet protein of unknown function. We show that it binds specifically to and is stabilized by droplets containing triacylglycerol (TG). Droplets isolated from cells with a PET10 deletion strongly aggregate, appear fragile, and fuse in vivo when cells are cultured in oleic acid. Pet10p binds early to nascent droplets, and their rate of appearance is decreased in pet10Δ. Moreover, Pet10p functionally interacts with the endoplasmic reticulum droplet assembly factors seipin and Fit2 to maintain proper droplet morphology. The activity of Dga1p, a diacylglycerol acyltransferase, and TG accumulation were both 30–35% lower in the absence of Pet10p. Pet10p contains a PAT domain, a defining property of perilipins, which was not previously known to exist in yeast. We propose that the core functions of Pet10p and other perilipins extend beyond protection from lipases and include the preservation of droplet integrity as well as collaboration with seipin and Fit2 in droplet assembly and maintenance. PMID:28801319

MESSI: metabolic engineering target selection and best strain identification tool.

PubMed

Kang, Kang; Li, Jun; Lim, Boon Leong; Panagiotou, Gianni

2015-01-01

Metabolic engineering and synthetic biology are synergistically related fields for manipulating target pathways and designing microorganisms that can act as chemical factories. Saccharomyces cerevisiae's ideal bioprocessing traits make yeast a very attractive chemical factory for production of fuels, pharmaceuticals, nutraceuticals as well as a wide range of chemicals. However, future attempts of engineering S. cerevisiae's metabolism using synthetic biology need to move towards more integrative models that incorporate the high connectivity of metabolic pathways and regulatory processes and the interactions in genetic elements across those pathways and processes. To contribute in this direction, we have developed Metabolic Engineering target Selection and best Strain Identification tool (MESSI), a web server for predicting efficient chassis and regulatory components for yeast bio-based production. The server provides an integrative platform for users to analyse ready-to-use public high-throughput metabolomic data, which are transformed to metabolic pathway activities for identifying the most efficient S. cerevisiae strain for the production of a compound of interest. As input MESSI accepts metabolite KEGG IDs or pathway names. MESSI outputs a ranked list of S. cerevisiae strains based on aggregation algorithms. Furthermore, through a genome-wide association study of the metabolic pathway activities with the strains' natural variation, MESSI prioritizes genes and small variants as potential regulatory points and promising metabolic engineering targets. Users can choose various parameters in the whole process such as (i) weight and expectation of each metabolic pathway activity in the final ranking of the strains, (ii) Weighted AddScore Fuse or Weighted Borda Fuse aggregation algorithm, (iii) type of variants to be included, (iv) variant sets in different biological levels.Database URL: http://sbb.hku.hk/MESSI/. © The Author(s) 2015. Published by Oxford University Press.

A model for cell wall dissolution in mating yeast cells: polarized secretion and restricted diffusion of cell wall remodeling enzymes induces local dissolution.

PubMed

Huberman, Lori B; Murray, Andrew W

2014-01-01

Mating of the budding yeast, Saccharomyces cerevisiae, occurs when two haploid cells of opposite mating types signal using reciprocal pheromones and receptors, grow towards each other, and fuse to form a single diploid cell. To fuse, both cells dissolve their cell walls at the point of contact. This event must be carefully controlled because the osmotic pressure differential between the cytoplasm and extracellular environment causes cells with unprotected plasma membranes to lyse. If the cell wall-degrading enzymes diffuse through the cell wall, their concentration would rise when two cells touched each other, such as when two pheromone-stimulated cells adhere to each other via mating agglutinins. At the surfaces that touch, the enzymes must diffuse laterally through the wall before they can escape into the medium, increasing the time the enzymes spend in the cell wall, and thus raising their concentration at the point of attachment and restricting cell wall dissolution to points where cells touch each other. We tested this hypothesis by studying pheromone treated cells confined between two solid, impermeable surfaces. This confinement increases the frequency of pheromone-induced cell death, and this effect is diminished by reducing the osmotic pressure difference across the cell wall or by deleting putative cell wall glucanases and other genes necessary for efficient cell wall fusion. Our results support the model that pheromone-induced cell death is the result of a contact-driven increase in the local concentration of cell wall remodeling enzymes and suggest that this process plays an important role in regulating cell wall dissolution and fusion in mating cells.

A Model for Cell Wall Dissolution in Mating Yeast Cells: Polarized Secretion and Restricted Diffusion of Cell Wall Remodeling Enzymes Induces Local Dissolution

PubMed Central

Huberman, Lori B.; Murray, Andrew W.

2014-01-01

Mating of the budding yeast, Saccharomyces cerevisiae, occurs when two haploid cells of opposite mating types signal using reciprocal pheromones and receptors, grow towards each other, and fuse to form a single diploid cell. To fuse, both cells dissolve their cell walls at the point of contact. This event must be carefully controlled because the osmotic pressure differential between the cytoplasm and extracellular environment causes cells with unprotected plasma membranes to lyse. If the cell wall-degrading enzymes diffuse through the cell wall, their concentration would rise when two cells touched each other, such as when two pheromone-stimulated cells adhere to each other via mating agglutinins. At the surfaces that touch, the enzymes must diffuse laterally through the wall before they can escape into the medium, increasing the time the enzymes spend in the cell wall, and thus raising their concentration at the point of attachment and restricting cell wall dissolution to points where cells touch each other. We tested this hypothesis by studying pheromone treated cells confined between two solid, impermeable surfaces. This confinement increases the frequency of pheromone-induced cell death, and this effect is diminished by reducing the osmotic pressure difference across the cell wall or by deleting putative cell wall glucanases and other genes necessary for efficient cell wall fusion. Our results support the model that pheromone-induced cell death is the result of a contact-driven increase in the local concentration of cell wall remodeling enzymes and suggest that this process plays an important role in regulating cell wall dissolution and fusion in mating cells. PMID:25329559

Multiplex PCR assay for detection of recombinant genes encoding fatty acid desaturases fused with lichenase reporter protein in GM plants.

PubMed

Berdichevets, Iryna N; Shimshilashvili, Hristina R; Gerasymenko, Iryna M; Sindarovska, Yana R; Sheludko, Yuriy V; Goldenkova-Pavlova, Irina V

2010-07-01

Thermostable lichenase encoded by licB gene of Clostridium thermocellum can be used as a reporter protein in plant, bacterial, yeast, and mammalian cells. It has important advantages of high sensitivity and specificity in qualitative and quantitative assays. Deletion variants of LicB (e.g., LicBM3) retain its enzymatic activity and thermostability and can be expressed in translational fusion with target proteins without compromising with their properties. Fusion with the lichenase reporter is especially convenient for the heterologous expression of proteins whose analysis is difficult or compromised by host enzyme activities, as it is in case of fatty acid desaturases occurring in all groups of organisms. Recombinant desaturase-lichenase genes can be used for creating genetically modified (GM) plants with improved chill tolerance. Development of an analytical method for detection of fused desaturase-lichenase transgenes is necessary both for production of GM plants and for their certification. Here, we report a multiplex polymerase chain reaction method for detection of desA and desC desaturase genes of cyanobacteria Synechocystis sp. PCC6803 and Synechococcus vulcanus, respectively, fused to licBM3 reporter in GM plants.

Assays for the biochemical and ultrastructural measurement of selective and nonselective types of autophagy in the yeast Saccharomyces cerevisiae.

PubMed

Guimaraes, Rodrigo Soares; Delorme-Axford, Elizabeth; Klionsky, Daniel J; Reggiori, Fulvio

2015-03-01

Autophagy is a conserved intracellular catabolic pathway that degrades unnecessary or dysfunctional cellular components. Components destined for degradation are sequestered into double-membrane vesicles called autophagosomes, which subsequently fuse with the vacuole/lysosome delivering their cargo into the interior of this organelle for turnover. Autophagosomes are generated through the concerted action of the autophagy-related (Atg) proteins. The yeast Saccharomyces cerevisiae has been key in the identification of the corresponding genes and their characterization, and it remains one of the leading model systems for the investigation of the molecular mechanism and functions of autophagy. In particular, it is still pivotal for the study of selective types of autophagy. The objective of this review is to present detailed protocols of the methods available to monitor the progression of both nonselective and selective types of autophagy, and to discuss their advantages and disadvantages. The ultimate aim is to provide researchers with the information necessary to select the optimal approach to address their biological question. Copyright © 2014 Elsevier Inc. All rights reserved.

Translation, modification and cellular distribution of two AC4 variants of African cassava mosaic virus in yeast and their pathogenic potential in plants

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hipp, Katharina, E-mail: katharina.hipp@bio.uni-st

Plant infecting geminiviruses encode a small (A)C4 protein within the open reading frame of the replication-initiator protein. In African cassava mosaic virus, two in-frame start codons may be used for the translation of a longer and a shorter AC4 variant. Both were fused to green fluorescent protein or glutathione-S-transferase genes and expressed in fission yeast. The longer variant accumulated in discrete spots in the cytoplasm, whereas the shorter variant localized to the plasma membrane. A similar expression pattern was found in plants. A myristoylation motif may promote a targeting of the shorter variant to the plasma membrane. Mass spectrometry analysismore » of the yeast-expressed shorter variant detected the corresponding myristoylation. The biological relevance of the second start codon was confirmed using mutated infectious clones. Whereas mutating the first start codon had no effect on the infectivity in Nicotiana benthamiana plants, the second start codon proved to be essential. -- Highlights: •The ACMV AC4 may be translated from one or the other in-frame start codon. •Both AC4 variants are translated in fission yeast. •The long AC4 protein localizes to the cytoplasm, the short to the plasma membrane. •The short variant is myristoylated in yeast and may promote membrane localization. •Only the shorter AC4 variant has an impact on viral infections in plants.« less

An insertion in the methyltransferase domain of P. falciparum trimethylguanosine synthase harbors a classical nuclear localization signal.

PubMed

Babar, Prasad H; Dey, Vishakha; Jaiswar, Praveen; Patankar, Swati

Many Plasmodium falciparum proteins do not share homology with, and are generally longer than their respective orthologs. This, to some extent, can be attributed to insertions. Here, we studied a P. falciparum RNA hypermethylase, trimethylguanosine synthase (PfTGS1) that harbors a 76 amino acid insertion in its methyltransferase domain. Bioinformatics analysis revealed that this insertion was present in TGS1 orthologs from other Plasmodium species as well. Interestingly, a classical nuclear localization signal (NLS) was predicted in the insertions of primate parasite TGS1 proteins. To check whether these predicted NLS are functional, we developed an in vivo heterologous system using S. cerevisiae. The predicted NLS when fused to dimeric GFP were able to localize the fusion protein to the nucleus in yeast indicating that it is indeed recognized by the yeast nuclear import machinery. We further showed that the PfTGS1 NLS binds to P. falciparum importin-α in vitro, confirming that the NLS is also recognized by the P. falciparum classical nuclear import machinery. Thus, in this study we report a novel function of the insertion in PfTGS1. Copyright © 2016 Elsevier B.V. All rights reserved.

Redesigning of Microbial Cell Surface and Its Application to Whole-Cell Biocatalysis and Biosensors.

PubMed

Han, Lei; Zhao, Yukun; Cui, Shan; Liang, Bo

2018-06-01

Microbial cell surface display technology can redesign cell surfaces with functional proteins and peptides to endow cells some unique features. Foreign peptides or proteins are transported out of cells and immobilized on cell surface by fusing with anchoring proteins, which is an effective solution to avoid substance transfer limitation, enzyme purification, and enzyme instability. As the most frequently used prokaryotic and eukaryotic protein surface display system, bacterial and yeast surface display systems have been widely applied in vaccine, biocatalysis, biosensor, bioadsorption, and polypeptide library screening. In this review of bacterial and yeast surface display systems, different cell surface display mechanisms and their applications in biocatalysis as well as biosensors are described with their strengths and shortcomings. In addition to single enzyme display systems, multi-enzyme co-display systems are presented here. Finally, future developments based on our and other previous reports are discussed.

Nuclear fusion and genome encounter during yeast zygote formation.

PubMed

Tartakoff, Alan Michael; Jaiswal, Purnima

2009-06-01

When haploid cells of Saccharomyces cerevisiae are crossed, parental nuclei congress and fuse with each other. To investigate underlying mechanisms, we have developed assays that evaluate the impact of drugs and mutations. Nuclear congression is inhibited by drugs that perturb the actin and tubulin cytoskeletons. Nuclear envelope (NE) fusion consists of at least five steps in which preliminary modifications are followed by controlled flux of first outer and then inner membrane proteins, all before visible dilation of the waist of the nucleus or coalescence of the parental spindle pole bodies. Flux of nuclear pore complexes occurs after dilation. Karyogamy requires both the Sec18p/NSF ATPase and ER/NE luminal homeostasis. After fusion, chromosome tethering keeps tagged parental genomes separate from each other. The process of NE fusion and evidence of genome independence in yeast provide a prototype for understanding related events in higher eukaryotes.

A general strategy to construct small molecule biosensors in eukaryotes.

PubMed

Feng, Justin; Jester, Benjamin W; Tinberg, Christine E; Mandell, Daniel J; Antunes, Mauricio S; Chari, Raj; Morey, Kevin J; Rios, Xavier; Medford, June I; Church, George M; Fields, Stanley; Baker, David

2015-12-29

Biosensors for small molecules can be used in applications that range from metabolic engineering to orthogonal control of transcription. Here, we produce biosensors based on a ligand-binding domain (LBD) by using a method that, in principle, can be applied to any target molecule. The LBD is fused to either a fluorescent protein or a transcriptional activator and is destabilized by mutation such that the fusion accumulates only in cells containing the target ligand. We illustrate the power of this method by developing biosensors for digoxin and progesterone. Addition of ligand to yeast, mammalian, or plant cells expressing a biosensor activates transcription with a dynamic range of up to ~100-fold. We use the biosensors to improve the biotransformation of pregnenolone to progesterone in yeast and to regulate CRISPR activity in mammalian cells. This work provides a general methodology to develop biosensors for a broad range of molecules in eukaryotes.

Arming Technology in Yeast-Novel Strategy for Whole-cell Biocatalyst and Protein Engineering.

PubMed

Kuroda, Kouichi; Ueda, Mitsuyoshi

2013-09-09

Cell surface display of proteins/peptides, in contrast to the conventional intracellular expression, has many attractive features. This arming technology is especially effective when yeasts are used as a host, because eukaryotic modifications that are often required for functional use can be added to the surface-displayed proteins/peptides. A part of various cell wall or plasma membrane proteins can be genetically fused to the proteins/peptides of interest to be displayed. This technology, leading to the generation of so-called "arming technology", can be employed for basic and applied research purposes. In this article, we describe various strategies for the construction of arming yeasts, and outline the diverse applications of this technology to industrial processes such as biofuel and chemical productions, pollutant removal, and health-related processes, including oral vaccines. In addition, arming technology is suitable for protein engineering and directed evolution through high-throughput screening that is made possible by the feature that proteins/peptides displayed on cell surface can be directly analyzed using intact cells without concentration and purification. Actually, novel proteins/peptides with improved or developed functions have been created, and development of diagnostic/therapeutic antibodies are likely to benefit from this powerful approach.

Suppressor Mutations for Presenilin 1 Familial Alzheimer Disease Mutants Modulate γ-Secretase Activities.

PubMed

Futai, Eugene; Osawa, Satoko; Cai, Tetsuo; Fujisawa, Tomoya; Ishiura, Shoichi; Tomita, Taisuke

2016-01-01

γ-Secretase is a multisubunit membrane protein complex containing presenilin (PS1) as a catalytic subunit. Familial Alzheimer disease (FAD) mutations within PS1 were analyzed in yeast cells artificially expressing membrane-bound substrate, amyloid precursor protein, or Notch fused to Gal4 transcriptional activator. The FAD mutations, L166P and G384A (Leu-166 to Pro and Gly-384 to Ala substitution, respectively), were loss-of-function in yeast. We identified five amino acid substitutions that suppress the FAD mutations. The cleavage of amyloid precursor protein or Notch was recovered by the secondary mutations. We also found that secondary mutations alone activated the γ-secretase activity. FAD mutants with suppressor mutations, L432M or S438P within TMD9 together with a missense mutation in the second or sixth loops, regained γ-secretase activity when introduced into presenilin null mouse fibroblasts. Notably, the cells with suppressor mutants produced a decreased amount of Aβ42, which is responsible for Alzheimer disease. These results indicate that the yeast system is useful to screen for mutations and chemicals that modulate γ-secretase activity. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Aym1, a mouse meiotic gene identified by virtue of its ability to activate early meiotic genes in the yeast Saccharomyces cerevisiae.

PubMed

Malcov, Mira; Cesarkas, Karen; Stelzer, Gil; Shalom, Sarah; Dicken, Yosef; Naor, Yaniv; Goldstein, Ronald S; Sagee, Shira; Kassir, Yona; Don, Jeremy

2004-12-01

Our understanding of the molecular mechanisms that operate during differentiation of mitotically dividing spermatogonia cells into spermatocytes lags way behind what is known about other differentiating systems. Given the evolutionary conservation of the meiotic process, we screened for mouse proteins that could specifically activate early meiotic promoters in Saccharomyces cerevisiae yeast cells, when fused to the Gal4 activation domain (Gal4AD). Our screen yielded the Aym1 gene that encodes a short peptide of 45 amino acids. We show that a Gal4AD-AYM1 fusion protein activates expression of reporter genes through the promoters of the early meiosis-specific genes IME2 and HOP1, and that this activation is dependent on the DNA-binding protein Ume6. Aym1 is transcribed predominantly in mouse primary spermatocytes and in gonads of female embryos undergoing the corresponding meiotic divisions. Aym1 immunolocalized to nuclei of primary spermatocytes and oocytes and to specific type A spermatogonia cells, suggesting it might play a role in the processes leading to meiotic competence. The potential functional relationship between AYM1 and yeast proteins that regulate expression of early meiotic genes is discussed.

Zea mays Taxilin Protein Negatively Regulates Opaque-2 Transcriptional Activity by Causing a Change in Its Sub-Cellular Distribution

PubMed Central

Zhang, Nan; Qiao, Zhenyi; Liang, Zheng; Mei, Bing; Xu, Zhengkai; Song, Rentao

2012-01-01

Zea mays (maize) Opaque-2 (ZmO2) protein is an important bZIP transcription factor that regulates the expression of major storage proteins (22-kD zeins) and other important genes during maize seed development. ZmO2 is subject to functional regulation through protein-protein interactions. To unveil the potential regulatory network associated with ZmO2, a protein-protein interaction study was carried out using the truncated version of ZmO2 (O2-2) as bait in a yeast two-hybrid screen with a maize seed cDNA library. A protein with homology to Taxilin was found to have stable interaction with ZmO2 in yeast and was designated as ZmTaxilin. Sequence analysis indicated that ZmTaxilin has a long coiled-coil domain containing three conserved zipper motifs. Each of the three zipper motifs is individually able to interact with ZmO2 in yeast. A GST pull-down assay demonstrated the interaction between GST-fused ZmTaxilin and ZmO2 extracted from developing maize seeds. Using onion epidermal cells as in vivo assay system, we found that ZmTaxilin could change the sub-cellular distribution of ZmO2. We also demonstrated that this change significantly repressed the transcriptional activity of ZmO2 on the 22-kD zein promoter. Our study suggests that a Taxilin-mediated change in sub-cellular distribution of ZmO2 may have important functional consequences for ZmO2 activity. PMID:22937104

Golgi Localized Barley MTP8 Proteins Facilitate Mn Transport

PubMed Central

Pedas, Pai; Schiller Stokholm, Michaela; Hegelund, Josefine Nymark; Ladegård, Anne Hald; Schjoerring, Jan Kofod; Husted, Søren

2014-01-01

Many metabolic processes in plants are regulated by manganese (Mn) but limited information is available on the molecular mechanisms controlling cellular Mn homeostasis. In this study, a yeast assay was used to isolate and characterize two genes, MTP8.1 and MTP8.2, which encode membrane-bound proteins belonging to the cation diffusion facilitator (CDF) family in the cereal species barley (Hordeum vulgare). Transient expression in onion epidermal cells showed that MTP8.1 and MTP8.2 proteins fused to the green fluorescent protein (GFP) are localized to Golgi. When heterologously expressed in yeast, MTP8.1 and MTP8.2 were found to be Mn transporters catalysing Mn efflux in a similar manner as the Golgi localized endogenous yeast protein Pmr1p. The level of MTP8.1 transcripts in barley roots increased with external Mn supply ranging from deficiency to toxicity, while MTP8.2 transcripts decreased under the same conditions, indicating non-overlapping functions for the two genes. In barley leaves, the expression of both MTP8 genes declined in response to toxic Mn additions to the roots suggesting a role in ensuring proper delivery of Mn to Golgi. Based on the above we suggest that barley MTP8 proteins are involved in Mn loading to the Golgi apparatus and play a role in Mn homeostasis by delivering Mn to Mn-dependent enzymes and/or by facilitating Mn efflux via secretory vesicles. This study highlights the importance of MTP transporters in Mn homeostasis and is the first report of Golgi localized Mn2+ transport proteins in a monocot plant species. PMID:25486417

Gal4-VP16 directs ATP-independent chromatin reorganization in a yeast chromatin assembly system.

PubMed

Robinson, Karen M; Schultz, Michael C

2005-03-22

Major insights into the regulation of chromatin organization have stemmed from biochemical studies using Gal4-VP16, a chimeric transcriptional activator in which the DNA binding domain of Gal4p is fused to the activation domain of viral protein VP16. Unexpectedly, given previous intensive efforts to understand how Gal4-VP16 functions in the context of chromatin, we have uncovered a new mode of chromatin reorganization that is dependent on Gal4-VP16. This reorganization is performed by an activity in a crude DEAE (CD) fraction from budding yeast which also supports ATP-dependent assembly of physiologically spaced nucleosome arrays. Biochemical analysis reveals that the activity tightly associates with chromatin and reorganizes nucleosome arrays by a mechanism which is insensitive to ATP depletion after nucleosome assembly. It generates a chromatin organization in which a nucleosome is stably positioned immediately adjacent to Gal4p binding sites in the template DNA. Individual deletion of genes previously implicated in chromatin assembly and remodeling, namely, the histone chaperones NAP1, ASF1, and CAC1 and the SNF2-like DEAD/H ATPases SNF2, ISW1, ISW2, CHD1, SWR1, YFR038w, and SPT20, does not significantly perturb reorganization. Therefore, Gal4-VP16-directed chromatin reorganization in yeast can occur by an ATP-independent mechanism that does not require SAGA, SWI/SNF, Isw1, or Isw2 chromatin remodeling complexes.

Improving genetic immobilization of a cellulase on yeast cell surface for bioethanol production using cellulose.

PubMed

Yang, Jinying; Dang, Hongyue; Lu, Jian Ren

2013-04-01

In this study, Saccharomyces cerevisiae was genetically engineered to harbor the capability of utilizing celluloses for bioethanol production by displaying active cellulolytic enzymes on the cell surface. An endo-1,4-β-glucanase gene egX was cloned from Bacillus pumilus C-9 and its expression products, the EGX cellulases, were displayed on the cell surface of S. cerevisiae by fusing egX with aga2 that encodes the binding subunit of the S. cerevisiae cell wall protein α-agglutinin. To achieve high gene copies and stability, multicopy integration was obtained by integrating the fusion aga2-egX gene into the rDNA region of the S. cerevisiae chromosome. To achieve high expression and surface display efficiency, the aga2-egX gene was expressed under the control of a strong promoter. The presence of the enzymatically active cellulase fusion proteins on the S. cerevisiae cell surface was verified by carboxymethyl cellulase activity assay and immunofluorescence microscopy. This work presented a promising strategy to genetically engineer yeasts to perform efficient fermentation of cellulosic materials for bioethanol production. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The First Prokaryotic Trehalose Synthase Complex Identified in the Hyperthermophilic Crenarchaeon Thermoproteus tenax

PubMed Central

Bräsen, Christopher; Hensel, Reinhard; Lupas, Andrei N.; Brinkmann, Henner; Siebers, Bettina

2013-01-01

The role of the disaccharide trehalose, its biosynthesis pathways and their regulation in Archaea are still ambiguous. In Thermoproteus tenax a fused trehalose-6-phosphate synthase/phosphatase (TPSP), consisting of an N-terminal trehalose-6-phosphate synthase (TPS) and a C-terminal trehalose-6-phosphate phosphatase (TPP) domain, was identified. The tpsp gene is organized in an operon with a putative glycosyltransferase (GT) and a putative mechanosensitive channel (MSC). The T. tenax TPSP exhibits high phosphatase activity, but requires activation by the co-expressed GT for bifunctional synthase-phosphatase activity. The GT mediated activation of TPS activity relies on the fusion of both, TPS and TPP domain, in the TPSP enzyme. Activation is mediated by complex-formation in vivo as indicated by yeast two-hybrid and crude extract analysis. In combination with first evidence for MSC activity the results suggest a sophisticated stress response involving TPSP, GT and MSC in T. tenax and probably in other Thermoproteales species. The monophyletic prokaryotic TPSP proteins likely originated via a single fusion event in the Bacteroidetes with subsequent horizontal gene transfers to other Bacteria and Archaea. Furthermore, evidence for the origin of eukaryotic TPSP fusions via HGT from prokaryotes and therefore a monophyletic origin of eukaryotic and prokaryotic fused TPSPs is presented. This is the first report of a prokaryotic, archaeal trehalose synthase complex exhibiting a much more simple composition than the eukaryotic complex described in yeast. Thus, complex formation and a complex-associated regulatory potential might represent a more general feature of trehalose synthesizing proteins. PMID:23626675

Purification and characterisation of the yeast plasma membrane ATP binding cassette transporter Pdr11p

PubMed Central

Laub, Katrine Rude; Marek, Magdalena; Stanchev, Lyubomir Dimitrov; Herrera, Sara Abad; Kanashova, Tamara; Bourmaud, Adèle; Dittmar, Gunnar

2017-01-01

The ATP binding cassette (ABC) transporters Pdr11p and its paralog Aus1p are expressed under anaerobic growth conditions at the plasma membrane of the yeast Saccharomyces cerevisiae and are required for sterol uptake. However, the precise mechanism by which these ABC transporters facilitate sterol movement is unknown. In this study, an overexpression and purification procedure was developed with the aim to characterise the Pdr11p transporter. Engineering of Pdr11p variants fused at the C terminus with green fluorescent protein (Pdr11p-GFP) and containing a FLAG tag at the N terminus facilitated expression analysis and one-step purification, respectively. The detergent-solubilised and purified protein displayed a stable ATPase activity with a broad pH optimum near 7.4. Mutagenesis of the conserved lysine to methionine (K788M) in the Walker A motif abolished ATP hydrolysis. Remarkably, and in contrast to Aus1p, ATPase activity of Pdr11p was insensitive to orthovanadate and not specifically stimulated by phosphatidylserine upon reconstitution into liposomes. Our results highlight distinct differences between Pdr11p and Aus1p and create an experimental basis for further biochemical studies of both ABC transporters to elucidate their function. PMID:28922409

Dominant genetics using a yeast genomic library under the control of a strong inducible promoter.

PubMed

Ramer, S W; Elledge, S J; Davis, R W

1992-12-01

In Saccharomyces cerevisiae, numerous genes have been identified by selection from high-copy-number libraries based on "multicopy suppression" or other phenotypic consequences of overexpression. Although fruitful, this approach suffers from two major drawbacks. First, high copy number alone may not permit high-level expression of tightly regulated genes. Conversely, other genes expressed in proportion to dosage cannot be identified if their products are toxic at elevated levels. This work reports construction of a genomic DNA expression library for S. cerevisiae that circumvents both limitations by fusing randomly sheared genomic DNA to the strong, inducible yeast GAL1 promoter, which can be regulated by carbon source. The library obtained contains 5 x 10(7) independent recombinants, representing a breakpoint at every base in the yeast genome. This library was used to examine aberrant gene expression in S. cerevisiae. A screen for dominant activators of yeast mating response identified eight genes that activate the pathway in the absence of exogenous mating pheromone, including one previously unidentified gene. One activator was a truncated STE11 gene lacking approximately 1000 base pairs of amino-terminal coding sequence. In two different clones, the same GAL1 promoter-proximal ATG is in-frame with the coding sequence of STE11, suggesting that internal initiation of translation there results in production of a biologically active, truncated STE11 protein. Thus this library allows isolation based on dominant phenotypes of genes that might have been difficult or impossible to isolate from high-copy-number libraries.

In-Vivo Real-Time Control of Protein Expression from Endogenous and Synthetic Gene Networks

PubMed Central

Orabona, Emanuele; De Stefano, Luca; Ferry, Mike; Hasty, Jeff; di Bernardo, Mario; di Bernardo, Diego

2014-01-01

We describe an innovative experimental and computational approach to control the expression of a protein in a population of yeast cells. We designed a simple control algorithm to automatically regulate the administration of inducer molecules to the cells by comparing the actual protein expression level in the cell population with the desired expression level. We then built an automated platform based on a microfluidic device, a time-lapse microscopy apparatus, and a set of motorized syringes, all controlled by a computer. We tested the platform to force yeast cells to express a desired fixed, or time-varying, amount of a reporter protein over thousands of minutes. The computer automatically switched the type of sugar administered to the cells, its concentration and its duration, according to the control algorithm. Our approach can be used to control expression of any protein, fused to a fluorescent reporter, provided that an external molecule known to (indirectly) affect its promoter activity is available. PMID:24831205

The yeast DNA ligase gene CDC9 is controlled by six orientation specific upstream activating sequences that respond to cellular proliferation but which alone cannot mediate cell cycle regulation.

PubMed Central

White, J H; Johnson, A L; Lowndes, N F; Johnston, L H

1991-01-01

By fusing the CDC9 structural gene to the PGK upstream sequences and the CDC9 upstream to lacZ, we showed that the cell cycle expression of CDC9 is largely due to transcriptional regulation. To investigate the role of six ATGATT upstream repeats in CDC9 regulation, synthetic copies of the sequence were attached to a heterologous gene. The repeats stimulated transcription strongly and additively, but, unlike conventional yeast UAS elements, only when present in one orientation. Transcription driven by the repeats declines in cells held at START of the cell cycle or in stationary phase, as occurs with CDC9. However, the repeats by themselves cannot impart cell cycle regulation to a heterologous gene. CDC9 may therefore be controlled by an activating system operating through the repeats that is sensitive to cellular proliferation and a separate mechanism that governs the periodic expression in the cell cycle. Images PMID:1901644

A general strategy to construct small molecule biosensors in eukaryotes

DOE PAGES

Feng, Justin; Jester, Benjamin W.; Tinberg, Christine E.; ...

2015-12-29

Biosensors for small molecules can be used in applications that range from metabolic engineering to orthogonal control of transcription. Here, we produce biosensors based on a ligand-binding domain (LBD) by using a method that, in principle, can be applied to any target molecule. The LBD is fused to either a fluorescent protein or a transcriptional activator and is destabilized by mutation such that the fusion accumulates only in cells containing the target ligand. We illustrate the power of this method by developing biosensors for digoxin and progesterone. Addition of ligand to yeast, mammalian, or plant cells expressing a biosensor activatesmore » transcription with a dynamic range of up to ~100-fold. We use the biosensors to improve the biotransformation of pregnenolone to progesterone in yeast and to regulate CRISPR activity in mammalian cells. As a result, this work provides a general methodology to develop biosensors for a broad range of molecules in eukaryotes.« less

A general strategy to construct small molecule biosensors in eukaryotes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Feng, Justin; Jester, Benjamin W.; Tinberg, Christine E.

Biosensors for small molecules can be used in applications that range from metabolic engineering to orthogonal control of transcription. Here, we produce biosensors based on a ligand-binding domain (LBD) by using a method that, in principle, can be applied to any target molecule. The LBD is fused to either a fluorescent protein or a transcriptional activator and is destabilized by mutation such that the fusion accumulates only in cells containing the target ligand. We illustrate the power of this method by developing biosensors for digoxin and progesterone. Addition of ligand to yeast, mammalian, or plant cells expressing a biosensor activatesmore » transcription with a dynamic range of up to ~100-fold. We use the biosensors to improve the biotransformation of pregnenolone to progesterone in yeast and to regulate CRISPR activity in mammalian cells. As a result, this work provides a general methodology to develop biosensors for a broad range of molecules in eukaryotes.« less

Matrix factorization-based data fusion for gene function prediction in baker's yeast and slime mold.

PubMed

Zitnik, Marinka; Zupan, Blaž

2014-01-01

The development of effective methods for the characterization of gene functions that are able to combine diverse data sources in a sound and easily-extendible way is an important goal in computational biology. We have previously developed a general matrix factorization-based data fusion approach for gene function prediction. In this manuscript, we show that this data fusion approach can be applied to gene function prediction and that it can fuse various heterogeneous data sources, such as gene expression profiles, known protein annotations, interaction and literature data. The fusion is achieved by simultaneous matrix tri-factorization that shares matrix factors between sources. We demonstrate the effectiveness of the approach by evaluating its performance on predicting ontological annotations in slime mold D. discoideum and on recognizing proteins of baker's yeast S. cerevisiae that participate in the ribosome or are located in the cell membrane. Our approach achieves predictive performance comparable to that of the state-of-the-art kernel-based data fusion, but requires fewer data preprocessing steps.

A general strategy to construct small molecule biosensors in eukaryotes

PubMed Central

Feng, Justin; Jester, Benjamin W; Tinberg, Christine E; Mandell, Daniel J; Antunes, Mauricio S; Chari, Raj; Morey, Kevin J; Rios, Xavier; Medford, June I; Church, George M; Fields, Stanley; Baker, David

2015-01-01

Biosensors for small molecules can be used in applications that range from metabolic engineering to orthogonal control of transcription. Here, we produce biosensors based on a ligand-binding domain (LBD) by using a method that, in principle, can be applied to any target molecule. The LBD is fused to either a fluorescent protein or a transcriptional activator and is destabilized by mutation such that the fusion accumulates only in cells containing the target ligand. We illustrate the power of this method by developing biosensors for digoxin and progesterone. Addition of ligand to yeast, mammalian, or plant cells expressing a biosensor activates transcription with a dynamic range of up to ~100-fold. We use the biosensors to improve the biotransformation of pregnenolone to progesterone in yeast and to regulate CRISPR activity in mammalian cells. This work provides a general methodology to develop biosensors for a broad range of molecules in eukaryotes. DOI: http://dx.doi.org/10.7554/eLife.10606.001 PMID:26714111

Cucumber metal transport protein MTP8 confers increased tolerance to manganese when expressed in yeast and Arabidopsis thaliana

PubMed Central

Migocka, Magdalena; Papierniak, Anna; Maciaszczyk-Dziubińska, Ewa; Poździk, Piotr; Posyniak, Ewelina; Garbiec, Arnold; Filleur, Sophie

2014-01-01

Cation diffusion facilitator (CDF) proteins are ubiquitous divalent cation transporters that have been proved to be essential for metal homeostasis and tolerance in Archaebacteria, Bacteria, and Eukaryota. In plants, CDFs are designated as metal tolerance proteins (MTPs). Due to the lack of genomic resources, studies on MTPs in other plants, including cultivated crops, are lacking. Here, the identification and organization of genes encoding members of the MTP family in cucumber are described. The first functional characterization of a cucumber gene encoding a member of the Mn-CDF subgroup of CDF proteins, designated as CsMTP8 based on the highest homology to plant MTP8, is also presented. The expression of CsMTP8 in Saccharomyces cerevisiae led to increased Mn accumulation in yeast cells and fully restored the growth of mutants hypersensitive to Mn in Mn excess. Similarly, the overexpression of CsMTP8 in Arabidopsis thaliana enhanced plant tolerance to high Mn in nutrition media as well as the accumulation of Mn in plant tissues. When fused to green fluorescent protein (GFP), CsMTP8 localized to the vacuolar membranes in yeast cells and to Arabidopsis protoplasts. In cucumber, CsMTP8 was expressed almost exclusively in roots, and the level of gene transcript was markedly up-regulated or reduced under elevated Mn or Mn deficiency, respectively. Taken together, the results suggest that CsMTP8 is an Mn transporter localized in the vacuolar membrane, which participates in the maintenance of Mn homeostasis in cucumber root cells. PMID:25039075

Development of a yeast heterologous expression cassette based on the promoter and terminator elements of the Eremothecium cymbalariae translational elongation factor 1α (EcTEF1) gene.

PubMed

Linder, Tomas

2018-04-01

A new expression cassette ( EC0 ) consisting of the fused 5' and 3' intergenic regions (IGRs) of the Eremothecium cymbalariae translational elongation factor 1α ( EcTEF1 ) gene was evaluated through expression of the bacterial hygromycin B phosphotransferase ( hph ) resistance gene in the common baker's yeast Saccharomyces cerevisiae . Progressively shorter versions of the hph -containing EC cassette ( hphEC1 though hphEC6 ) with trimmed 5' and 3' EcTEF1 IGRs were tested for their ability to confer resistance to hygromycin B in S. cerevisiae . Hygromycin B resistance was retained in all six generated hphEC variants up to a concentration of 400 mg/L. The hphEC6 cassette was the shortest cassette to be assayed in this study with 366 and 155 bp of the EcTEF1 5' and 3' IGRs, respectively. When tested for deletion of the S. cerevisiae proline oxidase gene PUT1 , the hphEC6 cassette was shown to successfully act as a selection marker on hygromycin B-containing medium. The hphEC6 cassette could be placed immediately adjacent to a kanMX4 G418 disulfate resistance marker without any discernable effect on the ability of the yeast to grow in the presence of both hygromycin B and G418 disulfate. Co-cultivation experiments under non-selective conditions demonstrated that a PUT1 deletion strain carrying the hphEC6 cassette displayed equivalent fitness to an otherwise isogenic PUT1 deletion strain carrying the kanMX4 cassette.

Structure and regulation of KGD1, the structural gene for yeast alpha-ketoglutarate dehydrogenase.

PubMed

Repetto, B; Tzagoloff, A

1989-06-01

Nuclear respiratory-defective mutants of Saccharomyces cerevisiae have been screened for lesions in the mitochondrial alpha-ketoglutarate dehydrogenase complex. Strains assigned to complementation group G70 were ascertained to be deficient in enzyme activity due to mutations in the KGD1 gene coding for the alpha-ketoglutarate dehydrogenase component of the complex. The KGD1 gene has been cloned by transformation of a representative kgd1 mutant, C225/U1, with a recombinant plasmid library of wild-type yeast nuclear DNA. Transformants containing the gene on a multicopy plasmid had three- to four-times-higher alpha-ketoglutarate dehydrogenase activity than did wild-type S. cerevisiae. Substitution of the chromosomal copy of KGD1 with a disrupted allele (kgd1::URA3) induced a deficiency in alpha-ketoglutarate dehydrogenase. The sequence of the cloned region of DNA which complements kgd1 mutants was found to have an open reading frame of 3,042 nucleotides capable of coding for a protein of Mw 114,470. The encoded protein had 38% identical residues with the reported sequence of alpha-ketoglutarate dehydrogenase from Escherichia coli. Two lines of evidence indicated that transcription of KGD1 is catabolite repressed. Higher steady-state levels of KGD1 mRNA were detected in wild-type yeast grown on the nonrepressible sugar galactose than in yeast grown on high glucose. Regulation of KGD1 was also studied by fusing different 5'-flanking regions of KGD1 to the lacZ gene of E. coli and measuring the expression of beta-galactosidase in yeast. Transformants harboring a fusion of 693 nucleotides of the 5'-flanking sequence expressed 10 times more beta-galactosidase activity when grown under derepressed conditions. The response to the carbon source was reduced dramatically when the same lacZ fusion was present in a hap2 or hap3 mutant. The promoter element(s) responsible for the regulated expression of KGD1 has been mapped to the -354 to -143 region. This region contained several putative activation sites with sequences matching the core element proposed to be essential for binding of the HAP2 and HAP3 regulatory proteins.

Characterizing the interactions between prolyl isomerase pin1 and phosphatase inhibitor-2 in living cells with FRET and FCS

NASA Astrophysics Data System (ADS)

Sun, Yuansheng; Wang, Lifu; Jyothikumar, Vinod; Brautigan, David L.; Periasamy, Ammasi

2012-03-01

Phosphatase inhibitor-2 (I2) was discovered as a regulator of protein Ser/Thr phosphatase-1 and is conserved from yeast to human. Binding between purified recombinant I2 from different species and the prolyl isomerase Pin1 has been demonstrated with pull-down assays, size exclusion chromatography and nuclear magnetic resonance spectroscopy. Despite this, questions persist as to whether these proteins associate together in living cells. In this study, we prepared fluorescent protein (FP) fusions of I2 and Pin1 and employed both Förster Resonance Energy Transfer (FRET) and Fluorescence Correlation Spectroscopy (FCS) imaging techniques to characterize their interactions in living cells. In both intensity-based and time-resolved FRET studies, we observed FRET uniformly across whole cells co-expressing I2-Cerulean and Pin1-Venus that was significantly higher than in negative controls expressing Cerulean FP (without fusing to I2) as the FRET donor and Pin1-Venus, showing a specific interaction between I2-Cerulean and Pin1-Venus in living cells. We also observed the co-diffusion of I2-Cerulean and Pin1-mCherry in Fluorescence Cross Correlation Spectroscopy (FCCS) measurements. We further showed that I2 itself as well as I2-Pin1 formed complexes in living cells (predicted from in vitro studies) via a quantitative FRET assay, and demonstrated from FCS measurements that both I2 and Pin1 (fused to Cerulean) are highly mobile in living cells.

The regulation of the SARK promoter activity by hormones and environmental signals.

PubMed

Delatorre, Carla A; Cohen, Yuval; Liu, Li; Peleg, Zvi; Blumwald, Eduardo

2012-09-01

The Senescence Associated Receptor Protein Kinase (P(SARK)) promoter, fused to isopentenyltransferase (IPT) gene has been shown to promote drought tolerance in crops. We dissected P(SARK) in order to understand the various elements associated with its activation and suppression. The activity of P(SARK) was higher in mature and early senescing leaves, and abiotic stress induced its activity in mature leaves. Bioinformatics analysis suggests the interactions of multiple cis-acting elements in the control of P(SARK) activity. In vitro gel shift assays and yeast one hybrid system revealed interactions of P(SARK) with transcription factors related to abscisic acid and cytokinin response. Deletion analysis of P(SARK), fused to GUS-reporter gene was used to identify specific regions regulating transcription under senescence or during drought stress. Effects of exogenous hormonal treatments were characterized in entire plants and in leaf disk assays, and regions responsive to various hormones were defined. Our results indicate a complex interaction of plant hormones and additional factors modulating P(SARK) activity under stress resulting in a transient induction of expression. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Garlic virus X 11-kDa protein granules move within the cytoplasm and traffic a host protein normally found in the nucleolus.

PubMed

Lu, Yuwen; Yan, Fei; Guo, Wei; Zheng, Hongying; Lin, Lin; Peng, Jiejun; Adams, Michael J; Chen, Jianping

2011-09-01

The subcellular localization of the 11-kDa protein (p11) encoded by ORF3 of Garlic virus X (GarVX; genus Allexivirus, family Alphaflexiviridae) was examined by confocal microscopy. Granules with intense fluorescence were visible on the endoplasmic reticulum when p11 fused with green or red fluorescent protein (GFP or RFP) was expressed in epidermal cells of Nicotiana benthamiana. Moreover, the p11-RFP granules moved in the cytoplasm, along the cell periphery and through the cell membranes to adjacent cells. A 17-kDa protein (p17) of garlic interacting with p11 was identified by yeast two-hybridization and bimolecular fluorescence complementation assay. When p17 fused to GFP was expressed in epidermal cells of N. benthamiana, it localized to the nucleolus. However, in the presence of GarVX p11, the distribution of p17 changed to that of p11, but did not appear to affect the pattern of movement of p11. MOLECULAR PLANT PATHOLOGY © 2011 BSPP AND BLACKWELL PUBLISHING LTD. NO CLAIM TO ORIGINAL US GOVERNMENT WORKS.

Identification of Candida glabrata genes involved in pH modulation and modification of the phagosomal environment in macrophages.

PubMed

Kasper, Lydia; Seider, Katja; Gerwien, Franziska; Allert, Stefanie; Brunke, Sascha; Schwarzmüller, Tobias; Ames, Lauren; Zubiria-Barrera, Cristina; Mansour, Michael K; Becken, Ulrike; Barz, Dagmar; Vyas, Jatin M; Reiling, Norbert; Haas, Albert; Haynes, Ken; Kuchler, Karl; Hube, Bernhard

2014-01-01

Candida glabrata currently ranks as the second most frequent cause of invasive candidiasis. Our previous work has shown that C. glabrata is adapted to intracellular survival in macrophages and replicates within non-acidified late endosomal-stage phagosomes. In contrast, heat killed yeasts are found in acidified matured phagosomes. In the present study, we aimed at elucidating the processes leading to inhibition of phagosome acidification and maturation. We show that phagosomes containing viable C. glabrata cells do not fuse with pre-labeled lysosomes and possess low phagosomal hydrolase activity. Inhibition of acidification occurs independent of macrophage type (human/murine), differentiation (M1-/M2-type) or activation status (vitamin D3 stimulation). We observed no differential activation of macrophage MAPK or NFκB signaling cascades downstream of pattern recognition receptors after internalization of viable compared to heat killed yeasts, but Syk activation decayed faster in macrophages containing viable yeasts. Thus, delivery of viable yeasts to non-matured phagosomes is likely not triggered by initial recognition events via MAPK or NFκB signaling, but Syk activation may be involved. Although V-ATPase is abundant in C. glabrata phagosomes, the influence of this proton pump on intracellular survival is low since blocking V-ATPase activity with bafilomycin A1 has no influence on fungal viability. Active pH modulation is one possible fungal strategy to change phagosome pH. In fact, C. glabrata is able to alkalinize its extracellular environment, when growing on amino acids as the sole carbon source in vitro. By screening a C. glabrata mutant library we identified genes important for environmental alkalinization that were further tested for their impact on phagosome pH. We found that the lack of fungal mannosyltransferases resulted in severely reduced alkalinization in vitro and in the delivery of C. glabrata to acidified phagosomes. Therefore, protein mannosylation may play a key role in alterations of phagosomal properties caused by C. glabrata.

Yor022c protein is a phospholipase A{sub 1} that localizes to the mitochondrial matrix

DOE Office of Scientific and Technical Information (OSTI.GOV)

Urafuji, Kyosei; Arioka, Manabu

In mammals, three types of intracellular phospholipase A{sub 1} (iPLA{sub 1}) enzymes have been characterized and are thought to be involved in various cellular processes such as phospholipid metabolism, organelle biogenesis, and membrane trafficking. In this study we analyzed the unique iPLA{sub 1}-like protein, Yor022c, in the budding yeast Saccharomyces cerevisiae. By the mass spectrometry analysis, we demonstrate that Yor022c is actually a phospholipase displaying sn-1-specific activity toward phosphatidylcholine, phosphatidylethanolamine, and phosphatidic acid, generating 2-acyl lysophospholipids. GFP-fused Yor022c co-stained with the mitochondrial dye MitoTracker, indicating that, unlike its mammalian counterparts, it is a mitochondrial protein. Further biochemical fractionation experiment combinedmore » with protease sensitivity assay showed that Yor022c localizes to the mitochondrial matrix. Thus Yor022c is the first PLA{sub 1} putatively involved in the maintenance of sn-1 acyl chains of phospholipids in the mitochondrial inner membrane. - Highlights: • Yeast Yor022c protein displays phospholipase A{sub 1} activity to various phospholipids. • Yor022c-GFP fusion protein localizes to mitochondria. • Biochemical fractionation showed that Yor022c localizes to the mitochondrial matrix.« less

Mapping DNA damage-dependent genetic interactions in yeast via party mating and barcode fusion genetics.

PubMed

Díaz-Mejía, J Javier; Celaj, Albi; Mellor, Joseph C; Coté, Atina; Balint, Attila; Ho, Brandon; Bansal, Pritpal; Shaeri, Fatemeh; Gebbia, Marinella; Weile, Jochen; Verby, Marta; Karkhanina, Anna; Zhang, YiFan; Wong, Cassandra; Rich, Justin; Prendergast, D'Arcy; Gupta, Gaurav; Öztürk, Sedide; Durocher, Daniel; Brown, Grant W; Roth, Frederick P

2018-05-28

Condition-dependent genetic interactions can reveal functional relationships between genes that are not evident under standard culture conditions. State-of-the-art yeast genetic interaction mapping, which relies on robotic manipulation of arrays of double-mutant strains, does not scale readily to multi-condition studies. Here, we describe barcode fusion genetics to map genetic interactions (BFG-GI), by which double-mutant strains generated via en masse "party" mating can also be monitored en masse for growth to detect genetic interactions. By using site-specific recombination to fuse two DNA barcodes, each representing a specific gene deletion, BFG-GI enables multiplexed quantitative tracking of double mutants via next-generation sequencing. We applied BFG-GI to a matrix of DNA repair genes under nine different conditions, including methyl methanesulfonate (MMS), 4-nitroquinoline 1-oxide (4NQO), bleomycin, zeocin, and three other DNA-damaging environments. BFG-GI recapitulated known genetic interactions and yielded new condition-dependent genetic interactions. We validated and further explored a subnetwork of condition-dependent genetic interactions involving MAG1 , SLX4, and genes encoding the Shu complex, and inferred that loss of the Shu complex leads to an increase in the activation of the checkpoint protein kinase Rad53. © 2018 The Authors. Published under the terms of the CC BY 4.0 license.

Screening for Glycosylphosphatidylinositol-Modified Cell Wall Proteins in Pichia pastoris and Their Recombinant Expression on the Cell Surface

PubMed Central

Zhang, Li; Liang, Shuli; Zhou, Xinying; Jin, Zi; Jiang, Fengchun; Han, Shuangyan; Zheng, Suiping

2013-01-01

Glycosylphosphatidylinositol (GPI)-anchored glycoproteins have various intrinsic functions in yeasts and different uses in vitro. In the present study, the genome of Pichia pastoris GS115 was screened for potential GPI-modified cell wall proteins. Fifty putative GPI-anchored proteins were selected on the basis of (i) the presence of a C-terminal GPI attachment signal sequence, (ii) the presence of an N-terminal signal sequence for secretion, and (iii) the absence of transmembrane domains in mature protein. The predicted GPI-anchored proteins were fused to an alpha-factor secretion signal as a substitute for their own N-terminal signal peptides and tagged with the chimeric reporters FLAG tag and mature Candida antarctica lipase B (CALB). The expression of fusion proteins on the cell surface of P. pastoris GS115 was determined by whole-cell flow cytometry and immunoblotting analysis of the cell wall extracts obtained by β-1,3-glucanase digestion. CALB displayed on the cell surface of P. pastoris GS115 with the predicted GPI-anchored proteins was examined on the basis of potential hydrolysis of p-nitrophenyl butyrate. Finally, 13 proteins were confirmed to be GPI-modified cell wall proteins in P. pastoris GS115, which can be used to display heterologous proteins on the yeast cell surface. PMID:23835174

Dynamic Assembly of Brambleberry Mediates Nuclear Envelope Fusion during Early Development

PubMed Central

Abrams, Elliott W.; Zhang, Hong; Marlow, Florence L.; Kapp, Lee; Lu, Sumei; Mullins, Mary C.

2012-01-01

Summary To accommodate the large cells following zygote formation, early blastomeres employ modified cell divisions. Karyomeres are one such modification, a mitotic intermediate wherein individual chromatin masses are surrounded by nuclear envelope, which then fuse to form a single mononucleus. We identified brambleberry, a maternal-effect zebrafish mutant that disrupts karyomere fusion resulting in formation of multiple micronuclei. brambleberry is a previously unannotated gene homologous to Kar5p, which participates in nuclear fusion in yeast. We demonstrate that Brambleberry is required for pronuclear fusion following fertilization in zebrafish. As karyomeres form, Brambleberry localizes to the nuclear envelope with prominent puncta evident near karyomere-karyomere interfaces corresponding to membrane fusion sites. Our studies identify the first factor acting in karyomere fusion and suggest that specialized proteins are necessary for proper nuclear division in large dividing blastomeres. PMID:22863006

Single molecule tracking fluorescence microscopy in mitochondria reveals highly dynamic but confined movement of Tom40

NASA Astrophysics Data System (ADS)

Kuzmenko, Anton; Tankov, Stoyan; English, Brian P.; Tarassov, Ivan; Tenson, Tanel; Kamenski, Piotr; Elf, Johan; Hauryliuk, Vasili

2011-12-01

Tom40 is an integral protein of the mitochondrial outer membrane, which as the central component of the Translocase of the Outer Membrane (TOM) complex forms a channel for protein import. We characterize the diffusion properties of individual Tom40 molecules fused to the photoconvertable fluorescent protein Dendra2 with millisecond temporal resolution. By imaging individual Tom40 molecules in intact isolated yeast mitochondria using photoactivated localization microscopy with sub-diffraction limited spatial precision, we demonstrate that Tom40 movement in the outer mitochondrial membrane is highly dynamic but confined in nature, suggesting anchoring of the TOM complex as a whole.

Deciphering the Origin, Evolution, and Physiological Function of the Subtelomeric Aryl-Alcohol Dehydrogenase Gene Family in the Yeast Saccharomyces cerevisiae.

PubMed

Yang, Dong-Dong; de Billerbeck, Gustavo M; Zhang, Jin-Jing; Rosenzweig, Frank; Francois, Jean-Marie

2018-01-01

Homology searches indicate that Saccharomyces cerevisiae strain BY4741 contains seven redundant genes that encode putative aryl-alcohol dehydrogenases (AAD). Yeast AAD genes are located in subtelomeric regions of different chromosomes, and their functional role(s) remain enigmatic. Here, we show that two of these genes, AAD4 and AAD14 , encode functional enzymes that reduce aliphatic and aryl-aldehydes concomitant with the oxidation of cofactor NADPH, and that Aad4p and Aad14p exhibit different substrate preference patterns. Other yeast AAD genes are undergoing pseudogenization. The 5' sequence of AAD15 has been deleted from the genome. Repair of an AAD3 missense mutation at the catalytically essential Tyr 73 residue did not result in a functional enzyme. However, ancestral-state reconstruction by fusing Aad6 with Aad16 and by N-terminal repair of Aad10 restores NADPH-dependent aryl-alcohol dehydrogenase activities. Phylogenetic analysis indicates that AAD genes are narrowly distributed in wood-saprophyte fungi and in yeast that occupy lignocellulosic niches. Because yeast AAD genes exhibit activity on veratraldehyde, cinnamaldehyde, and vanillin, they could serve to detoxify aryl-aldehydes released during lignin degradation. However, none of these compounds induce yeast AAD gene expression, and Aad activities do not relieve aryl-aldehyde growth inhibition. Our data suggest an ancestral role for AAD genes in lignin degradation that is degenerating as a result of yeast's domestication and use in brewing, baking, and other industrial applications. IMPORTANCE Functional characterization of hypothetical genes remains one of the chief tasks of the postgenomic era. Although the first Saccharomyces cerevisiae genome sequence was published over 20 years ago, 22% of its estimated 6,603 open reading frames (ORFs) remain unverified. One outstanding example of this category of genes is the enigmatic seven-member AAD family. Here, we demonstrate that proteins encoded by two members of this family exhibit aliphatic and aryl-aldehyde reductase activity, and further that such activity can be recovered from pseudogenized AAD genes via ancestral-state reconstruction. The phylogeny of yeast AAD genes suggests that these proteins may have played an important ancestral role in detoxifying aromatic aldehydes in ligninolytic fungi. However, in yeast adapted to niches rich in sugars, AAD genes become subject to mutational erosion. Our findings shed new light on the selective pressures and molecular mechanisms by which genes undergo pseudogenization. Copyright © 2017 Yang et al.

Deciphering the Origin, Evolution, and Physiological Function of the Subtelomeric Aryl-Alcohol Dehydrogenase Gene Family in the Yeast Saccharomyces cerevisiae

PubMed Central

de Billerbeck, Gustavo M.; Zhang, Jin-jing; Rosenzweig, Frank

2017-01-01

ABSTRACT Homology searches indicate that Saccharomyces cerevisiae strain BY4741 contains seven redundant genes that encode putative aryl-alcohol dehydrogenases (AAD). Yeast AAD genes are located in subtelomeric regions of different chromosomes, and their functional role(s) remain enigmatic. Here, we show that two of these genes, AAD4 and AAD14, encode functional enzymes that reduce aliphatic and aryl-aldehydes concomitant with the oxidation of cofactor NADPH, and that Aad4p and Aad14p exhibit different substrate preference patterns. Other yeast AAD genes are undergoing pseudogenization. The 5′ sequence of AAD15 has been deleted from the genome. Repair of an AAD3 missense mutation at the catalytically essential Tyr73 residue did not result in a functional enzyme. However, ancestral-state reconstruction by fusing Aad6 with Aad16 and by N-terminal repair of Aad10 restores NADPH-dependent aryl-alcohol dehydrogenase activities. Phylogenetic analysis indicates that AAD genes are narrowly distributed in wood-saprophyte fungi and in yeast that occupy lignocellulosic niches. Because yeast AAD genes exhibit activity on veratraldehyde, cinnamaldehyde, and vanillin, they could serve to detoxify aryl-aldehydes released during lignin degradation. However, none of these compounds induce yeast AAD gene expression, and Aad activities do not relieve aryl-aldehyde growth inhibition. Our data suggest an ancestral role for AAD genes in lignin degradation that is degenerating as a result of yeast's domestication and use in brewing, baking, and other industrial applications. IMPORTANCE Functional characterization of hypothetical genes remains one of the chief tasks of the postgenomic era. Although the first Saccharomyces cerevisiae genome sequence was published over 20 years ago, 22% of its estimated 6,603 open reading frames (ORFs) remain unverified. One outstanding example of this category of genes is the enigmatic seven-member AAD family. Here, we demonstrate that proteins encoded by two members of this family exhibit aliphatic and aryl-aldehyde reductase activity, and further that such activity can be recovered from pseudogenized AAD genes via ancestral-state reconstruction. The phylogeny of yeast AAD genes suggests that these proteins may have played an important ancestral role in detoxifying aromatic aldehydes in ligninolytic fungi. However, in yeast adapted to niches rich in sugars, AAD genes become subject to mutational erosion. Our findings shed new light on the selective pressures and molecular mechanisms by which genes undergo pseudogenization. PMID:29079624

Metabolic engineering for high glycerol production by the anaerobic cultures of Saccharomyces cerevisiae.

PubMed

Semkiv, Marta V; Dmytruk, Kostyantyn V; Abbas, Charles A; Sibirny, Andriy A

2017-06-01

Glycerol is used by the cosmetic, paint, automotive, food, and pharmaceutical industries and for production of explosives. Currently, glycerol is available in commercial quantities as a by-product from biodiesel production, but the purity and the cost of its purification are prohibitive. The industrial production of glycerol by glucose aerobic fermentation using osmotolerant strains of the yeasts Candida sp. and Saccharomyces cerevisiae has been described. A major drawback of the aerobic process is the high cost of production. For this reason, the development of yeast strains that effectively convert glucose to glycerol anaerobically is of great importance. Due to its ability to grow under anaerobic conditions, the yeast S. cerevisiae is an ideal system for the development of this new biotechnological platform. To increase glycerol production and accumulation from glucose, we lowered the expression of TPI1 gene coding for triose phosphate isomerase; overexpressed the fused gene consisting the GPD1 and GPP2 parts coding for glycerol-3-phosphate dehydrogenase and glycerol-3-phosphate phosphatase, respectively; overexpressed the engineered FPS1 gene that codes for aquaglyceroporin; and overexpressed the truncated gene ILV2 that codes for acetolactate synthase. The best constructed strain produced more than 20 g of glycerol/L from glucose under micro-aerobic conditions and 16 g of glycerol/L under anaerobic conditions. The increase in glycerol production led to a drop in ethanol and biomass accumulation.

MATRIX FACTORIZATION-BASED DATA FUSION FOR GENE FUNCTION PREDICTION IN BAKER’S YEAST AND SLIME MOLD

PubMed Central

ŽITNIK, MARINKA; ZUPAN, BLAŽ

2014-01-01

The development of effective methods for the characterization of gene functions that are able to combine diverse data sources in a sound and easily-extendible way is an important goal in computational biology. We have previously developed a general matrix factorization-based data fusion approach for gene function prediction. In this manuscript, we show that this data fusion approach can be applied to gene function prediction and that it can fuse various heterogeneous data sources, such as gene expression profiles, known protein annotations, interaction and literature data. The fusion is achieved by simultaneous matrix tri-factorization that shares matrix factors between sources. We demonstrate the effectiveness of the approach by evaluating its performance on predicting ontological annotations in slime mold D. discoideum and on recognizing proteins of baker’s yeast S. cerevisiae that participate in the ribosome or are located in the cell membrane. Our approach achieves predictive performance comparable to that of the state-of-the-art kernel-based data fusion, but requires fewer data preprocessing steps. PMID:24297565

Analysis of the 5′ untranslated region (5′UTR) of the alcohol oxidase 1 (AOX1) gene in recombinant protein expression in Pichia pastoris

PubMed Central

Staley, Chris A.; Huang, Amy; Nattestad, Maria; Oshiro, Kristin T.; Ray, Laura E.; Mulye, Tejas; Li, Zhiguo Harry; Le, Thu; Stephens, Justin J.; Gomez, Seth R.; Moy, Allison D.; Nguyen, Jackson C.; Franz, Andreas H.; Lin-Cereghino, Joan; Lin-Cereghino, Geoff P.

2012-01-01

Pichia pastoris is a methylotrophic yeast that has been genetically engineered to express over one thousand heterologous proteins valued for industrial, pharmaceutical and basic research purposes. In most cases, the 5′ untranslated region (UTR) of the alcohol oxidase 1 (AOX1) gene is fused to the coding sequence of the recombinant gene for protein expression in this yeast. Because the effect of the AOX1 5′UTR on protein expression is not known, site-directed mutagenesis was performed in order to decrease or increase the length of this region. Both of these types of changes were shown to affect translational efficiency, not transcript stability. While increasing the length of the 5′UTR clearly decreased expression of a β-galactosidase reporter in a proportional manner, a deletion analysis demonstrated that the AOX1 5′UTR contains a complex mixture of both positive and negative cis-acting elements, suggesting that the construction of a synthetic 5′UTR optimized for a higher level of expression may be challenging. PMID:22285974

Regulation of Compound Leaf Development in Medicago truncatula by Fused Compound Leaf1, a Class M KNOX Gene[C][W

PubMed Central

Peng, Jianling; Yu, Jianbin; Wang, Hongliang; Guo, Yingqing; Li, Guangming; Bai, Guihua; Chen, Rujin

2011-01-01

Medicago truncatula is a legume species belonging to the inverted repeat lacking clade (IRLC) with trifoliolate compound leaves. However, the regulatory mechanisms underlying development of trifoliolate leaves in legumes remain largely unknown. Here, we report isolation and characterization of fused compound leaf1 (fcl1) mutants of M. truncatula. Phenotypic analysis suggests that FCL1 plays a positive role in boundary separation and proximal-distal axis development of compound leaves. Map-based cloning indicates that FCL1 encodes a class M KNOX protein that harbors the MEINOX domain but lacks the homeodomain. Yeast two-hybrid assays show that FCL1 interacts with a subset of Arabidopsis thaliana BEL1-like proteins with slightly different substrate specificities from the Arabidopsis homolog KNATM-B. Double mutant analyses with M. truncatula single leaflet1 (sgl1) and palmate-like pentafoliata1 (palm1) leaf mutants show that fcl1 is epistatic to palm1 and sgl1 is epistatic to fcl1 in terms of leaf complexity and that SGL1 and FCL1 act additively and are required for petiole development. Previous studies have shown that the canonical KNOX proteins are not involved in compound leaf development in IRLC legumes. The identification of FCL1 supports the role of a truncated KNOX protein in compound leaf development in M. truncatula. PMID:22080596

Functional analysis of the isoforms of an ABI3-like factor of Pisum sativum generated by alternative splicing.

PubMed

Gagete, Andrés P; Riera, Marta; Franco, Luis; Rodrigo, M Isabel

2009-01-01

At least seven isoforms (PsABI3-1 to PsABI3-7) of a putative, pea ABI3-like factor, originated by alternative splicing, have been identified after cDNA cloning. A similar variability had previously only been described for monocot genes. The full-length isoform, PsABI3-1, contains the typical N-terminal acidic domains and C-terminal basic subdomains, B1 to B3. Reverse transcriptase-PCR analysis revealed that the gene is expressed just in seeds, starting at middle embryogenesis; no gene products are observed in embryo axes after 18 h post-imbibition although they are more persistent in cotyledons. The activity of the isoforms was studied by yeast one-hybrid assays. When yeast was transformed with the isoforms fused to the DNA binding domain of Gal4p, only the polypeptides PsABI3-2 and PsABI3-7 failed to complement the activity of Gal4p. Acidic domains A1 and A2 exhibit transactivating activity, but the former requires a small C-terminal extension to be active. Yeast two-hybrid analysis showed that PsABI3 is able to heterodimerize with Arabidopsis thaliana ABI5, thus proving that PsABI3 is functionally active. The minimum requirement for the interaction PsABI3-AtABI5 is the presence of the subdomain B1 with an extension, 81 amino acids long, at their C-terminal side. Finally, a transient onion transformation assay showed that both the active PsABI3-1 and the inactive PsABI3-2 isoforms are localized to nuclei. Considering that the major isoforms remain approximately constant in developing seeds although their relative proportion varied, the possible role of splicing in the regulatory network of ABA signalling is discussed.

Cassette Series Designed for Live-Cell Imaging of Proteins and High Resolution Techniques in Yeast

PubMed Central

Young, Carissa L.; Raden, David L.; Caplan, Jeffrey; Czymmek, Kirk; Robinson, Anne S.

2012-01-01

During the past decade, it has become clear that protein function and regulation are highly dependent upon intracellular localization. Although fluorescent protein variants are ubiquitously used to monitor protein dynamics, localization, and abundance; fluorescent light microscopy techniques often lack the resolution to explore protein heterogeneity and cellular ultrastructure. Several approaches have been developed to identify, characterize, and monitor the spatial localization of proteins and complexes at the sub-organelle level; yet, many of these techniques have not been applied to yeast. Thus, we have constructed a series of cassettes containing codon-optimized epitope tags, fluorescent protein variants that cover the full spectrum of visible light, a TetCys motif used for FlAsH-based localization, and the first evaluation in yeast of a photoswitchable variant – mEos2 – to monitor discrete subpopulations of proteins via confocal microscopy. This series of modules, complete with six different selection markers, provides the optimal flexibility during live-cell imaging and multicolor labeling in vivo. Furthermore, high-resolution imaging techniques include the yeast-enhanced TetCys motif that is compatible with diaminobenzidine photooxidation used for protein localization by electron microscopy and mEos2 that is ideal for super-resolution microscopy. We have examined the utility of our cassettes by analyzing all probes fused to the C-terminus of Sec61, a polytopic membrane protein of the endoplasmic reticulum of moderate protein concentration, in order to directly compare fluorescent probes, their utility and technical applications. Our series of cassettes expand the repertoire of molecular tools available to advance targeted spatiotemporal investigations using multiple live-cell, super-resolution or electron microscopy imaging techniques. PMID:22473760

Cell fusion and nuclear fusion in plants.

PubMed

Maruyama, Daisuke; Ohtsu, Mina; Higashiyama, Tetsuya

2016-12-01

Eukaryotic cells are surrounded by a plasma membrane and have a large nucleus containing the genomic DNA, which is enclosed by a nuclear envelope consisting of the outer and inner nuclear membranes. Although these membranes maintain the identity of cells, they sometimes fuse to each other, such as to produce a zygote during sexual reproduction or to give rise to other characteristically polyploid tissues. Recent studies have demonstrated that the mechanisms of plasma membrane or nuclear membrane fusion in plants are shared to some extent with those of yeasts and animals, despite the unique features of plant cells including thick cell walls and intercellular connections. Here, we summarize the key factors in the fusion of these membranes during plant reproduction, and also focus on "non-gametic cell fusion," which was thought to be rare in plant tissue, in which each cell is separated by a cell wall. Copyright © 2016 Elsevier Ltd. All rights reserved.

Dynamic assembly of brambleberry mediates nuclear envelope fusion during early development.

PubMed

Abrams, Elliott W; Zhang, Hong; Marlow, Florence L; Kapp, Lee; Lu, Sumei; Mullins, Mary C

2012-08-03

To accommodate the large cells following zygote formation, early blastomeres employ modified cell divisions. Karyomeres are one such modification, mitotic intermediates wherein individual chromatin masses are surrounded by nuclear envelope; the karyomeres then fuse to form a single mononucleus. We identified brambleberry, a maternal-effect zebrafish mutant that disrupts karyomere fusion, resulting in formation of multiple micronuclei. As karyomeres form, Brambleberry protein localizes to the nuclear envelope, with prominent puncta evident near karyomere-karyomere interfaces corresponding to membrane fusion sites. brambleberry corresponds to an unannotated gene with similarity to Kar5p, a protein that participates in nuclear fusion in yeast. We also demonstrate that Brambleberry is required for pronuclear fusion following fertilization in zebrafish. Our studies provide insight into the machinery required for karyomere fusion and suggest that specialized proteins are necessary for proper nuclear division in large dividing blastomeres. Copyright © 2012 Elsevier Inc. All rights reserved.

Interactions and Nuclear Import of the N and P Proteins of Sonchus Yellow Net Virus, a Plant Nucleorhabdovirus

PubMed Central

Goodin, Michael M.; Austin, Jennifer; Tobias, Renée; Fujita, Miki; Morales, Christina; Jackson, Andrew O.

2001-01-01

We have characterized the interaction and nuclear localization of the nucleocapsid (N) protein and phosphoprotein (P) of sonchus yellow net nucleorhabdovirus. Expression studies with plant and yeast cells revealed that both N and P are capable of independent nuclear import. Site-specific mutagenesis and deletion analyses demonstrated that N contains a carboxy-terminal bipartite nuclear localization signal (NLS) located between amino acids 465 and 481 and that P contains a karyophillic region between amino acids 40 and 124. The N NLS was fully capable of functioning outside of the context of the N protein and was able to direct the nuclear import of a synthetic protein fusion consisting of green fluorescent protein fused to glutathione S-transferase (GST). Expression and mapping studies suggested that the karyophillic domain in P is located within the N-binding domain. Coexpression of N and P drastically affected their localization patterns relative to those of individually expressed proteins and resulted in a shift of both proteins to a subnuclear region. Yeast two-hybrid and GST pulldown experiments verified the N-P and P-P interactions, and deletion analyses have identified the N and P interacting domains. N NLS mutants were not transported to the nucleus by import-competent P, presumably because N binding masks the P NLS. Taken together, our results support a model for independent entry of N and P into the nucleus followed by associations that mediate subnuclear localization. PMID:11533202

From fission to fusion: a perspective on the research that won the Nobel Prize in Physiology or Medicine, 2013.

PubMed

Ray, Krishanu

2014-03-01

Secretion is widespread in all eukaryotic cells: all of us experience this in the course of daily life--saliva, mucus, sweat, tears, bile juice, adrenalin, etc.--the list is extremely long. How does a cell manage to repeatedly spit out some stuff without losing the rest? The answer is: through regulated vesicle trafficking within the cell. The Nobel Prize in Physiology or Medicine 2013 was awarded to Drs Randy Schekman, James E Rothman and Thomas C Südhof for their 'discoveries of machinery regulating vesicle traffic, a major transport system in our cells'. Dr Randy Schekman and his colleagues discovered a number of genes required for vesicle trafficking from the endoplasmic reticulum (ER) and Golgi; the James E Rothman group unravelled the protein machinery that allows vesicles to bud off from the membrane and fuse to their targets; and Dr Thomas C Südhof along with his colleagues revealed how calcium ions could instruct vesicles to fuse and discharge their contents with precision. These enabled the biotechnology industry to produce a variety of pharmaceutical and industrial products like insulin and hepatitis B vaccines, in a cost-efficient manner, using yeast and tissue cultured cells.

Prions on the run: How extracellular vesicles serve as delivery vehicles for self-templating protein aggregates.

PubMed

Liu, Shu; Hossinger, André; Göbbels, Sarah; Vorberg, Ina M

2017-03-04

Extracellular vesicles (EVs) are actively secreted, membrane-bound communication vehicles that exchange biomolecules between cells. EVs also serve as dissemination vehicles for pathogens, including prions, proteinaceous infectious agents that cause transmissible spongiform encephalopathies (TSEs) in mammals. Increasing evidence accumulates that diverse protein aggregates associated with common neurodegenerative diseases are packaged into EVs as well. Vesicle-mediated intercellular transmission of protein aggregates can induce aggregation of homotypic proteins in acceptor cells and might thereby contribute to disease progression. Our knowledge of how protein aggregates are sorted into EVs and how these vesicles adhere to and fuse with target cells is limited. Here we review how TSE prions exploit EVs for intercellular transmission and compare this to the transmission behavior of self-templating cytosolic protein aggregates derived from the yeast prion domain Sup 35 NM. Artificial NM prions are non-toxic to mammalian cell cultures and do not cause loss-of-function phenotypes. Importantly, NM particles are also secreted in association with exosomes that horizontally transmit the prion phenotype to naive bystander cells, a process that can be monitored with high accuracy by automated high throughput confocal microscopy. The high abundance of mammalian proteins with amino acid stretches compositionally similar to yeast prion domains makes the NM cell model an attractive model to study self-templating and dissemination properties of proteins with prion-like domains in the mammalian context.

A visual screen for diet-regulated proteins in the Drosophila ovary using GFP protein trap lines

PubMed Central

Hsu, Hwei-Jan; Drummond-Barbosa, Daniela

2017-01-01

The effect of diet on reproduction is well documented in a large number of organisms; however, much remains to be learned about the molecular mechanisms underlying this connection. The Drosophila ovary has a well described, fast and largely reversible response to diet. Ovarian stem cells and their progeny proliferate and grow faster on a yeast-rich diet than on a yeast-free (poor) diet, and death of early germline cysts, degeneration of early vitellogenic follicles and partial block in ovulation further contribute to the ~60-fold decrease in egg laying observed on a poor diet. Multiple diet-dependent factors, including insulin-like peptides, the steroid ecdysone, the nutrient sensor Target of Rapamycin, AMP-dependent kinase, and adipocyte factors mediate this complex response. Here, we describe the results of a visual screen using a library of green fluorescent protein (GFP) protein trap lines to identify additional factors potentially involved in this response. In each GFP protein trap line, an artificial GFP exon is fused in frame to an endogenous protein, such that the GFP fusion pattern parallels the levels and subcellular localization of the corresponding native protein. We identified 53 GFP-tagged proteins that exhibit changes in levels and/or subcellular localization in the ovary at 12-16 hours after switching females from rich to poor diets, suggesting them as potential candidates for future functional studies. PMID:28093350

[Surface display of phytase on Saccharomyces cerevisiae for efficient bioethanol production from corn starch].

PubMed

Xiao, Yan; Chen, Xianzhong; Shen, Wei; Yang, Haiquan; Fan, You

2015-12-01

Production of bioethanol using starch as raw material has become a very prominent technology. However, phytate in the raw material not only decreases ethanol production efficiency, but also increases phosphorus discharge. In this study, to decrease phytate content in an ethanol fermentationprocess, Saccharomyces cerevisiae was engineered forheterologous expression of phytase on the cell surface. The phy gene encoding phytase gene was fused with the C-terminal-half region of α-agglutinin and then inserted downstream of the secretion signal gene, to produce a yeast surface-display expression vector pMGK-AG-phy, which was then transformed into S. cerevisiae. The recombinant yeast strain, PHY, successfully displayed phytase on the surface of cells producing 6.4 U/g wet cells and its properties were further characterized. The growthrate and ethanol production of the PHY strain were faster than the parent S. cerevisiae strain in the fermentation medium by simultaneous saccharification and fermentation. Moreover, the phytate concentration decreased by 91% in dry vinasse compared to the control. In summary, we constructed recombinant S. cerevisiae strain displaying phytase on the cell surface, which could effectively reduce the content of phytate, improve the utilization value of vinasse and reduce the discharge of phosphorus. The strain reported here represents a useful novel engineering platform for developing an environment-friendly system for bioethanol production from a corn substrate.

Enzyme reversal to explore the function of yeast E3 ubiquitin-ligases.

PubMed

MacDonald, Chris; Winistorfer, Stanley; Pope, Robert M; Wright, Michael E; Piper, Robert C

2017-07-01

The covalent attachment of ubiquitin onto proteins can elicit a variety of downstream consequences. Attachment is mediated by a large array of E3 ubiquitin ligases, each thought be subject to regulatory control and to have a specific repertoire of substrates. Assessing the biological roles of ligases, and in particular, identifying their biologically relevant substrates has been a persistent yet challenging question. In this study, we describe tools that may help achieve both of these goals. We describe a strategy whereby the activity of a ubiquitin ligase has been enzymatically reversed, accomplished by fusing it to a catalytic domain of an exogenous deubiquitinating enzyme. We present a library of 72 "anti-ligases" that appear to work in a dominant-negative fashion to stabilize their cognate substrates against ubiquitin-dependent proteasomal and lysosomal degradation. We then used the ligase-deubiquitinating enzyme (DUb) library to screen for E3 ligases involved in post-Golgi/endosomal trafficking. We identify ligases previously implicated in these pathways (Rsp5 and Tul1), in addition to ligases previously localized to endosomes (Pib1 and Vps8). We also document an optimized workflow for isolating and analyzing the "ubiquitome" of yeast, which can be used with mass spectrometry to identify substrates perturbed by expression of particular ligase-DUb fusions. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Mitochondrial anchorage and fusion contribute to mitochondrial inheritance and quality control in the budding yeast Saccharomyces cerevisiae

PubMed Central

Higuchi-Sanabria, Ryo; Charalel, Joseph K.; Viana, Matheus P.; Garcia, Enrique J.; Sing, Cierra N.; Koenigsberg, Andrea; Swayne, Theresa C.; Vevea, Jason D.; Boldogh, Istvan R.; Rafelski, Susanne M.; Pon, Liza A.

2016-01-01

Higher-functioning mitochondria that are more reduced and have less ROS are anchored in the yeast bud tip by the Dsl1-family protein Mmr1p. Here we report a role for mitochondrial fusion in bud-tip anchorage of mitochondria. Fluorescence loss in photobleaching (FLIP) and network analysis experiments revealed that mitochondria in large buds are a continuous reticulum that is physically distinct from mitochondria in mother cells. FLIP studies also showed that mitochondria that enter the bud can fuse with mitochondria that are anchored in the bud tip. In addition, loss of fusion and mitochondrial DNA (mtDNA) by deletion of mitochondrial outer or inner membrane fusion proteins (Fzo1p or Mgm1p) leads to decreased accumulation of mitochondria at the bud tip and inheritance of fitter mitochondria by buds compared with cells with no mtDNA. Conversely, increasing the accumulation and anchorage of mitochondria in the bud tip by overexpression of MMR1 results in inheritance of less-fit mitochondria by buds and decreased replicative lifespan and healthspan. Thus quantity and quality of mitochondrial inheritance are ensured by two opposing processes: bud-tip anchorage by mitochondrial fusion and Mmr1p, which favors bulk inheritance; and quality control mechanisms that promote segregation of fitter mitochondria to the bud. PMID:26764088

Inferring gene expression from ribosomal promoter sequences, a crowdsourcing approach

PubMed Central

Meyer, Pablo; Siwo, Geoffrey; Zeevi, Danny; Sharon, Eilon; Norel, Raquel; Segal, Eran; Stolovitzky, Gustavo; Siwo, Geoffrey; Rider, Andrew K.; Tan, Asako; Pinapati, Richard S.; Emrich, Scott; Chawla, Nitesh; Ferdig, Michael T.; Tung, Yi-An; Chen, Yong-Syuan; Chen, Mei-Ju May; Chen, Chien-Yu; Knight, Jason M.; Sahraeian, Sayed Mohammad Ebrahim; Esfahani, Mohammad Shahrokh; Dreos, Rene; Bucher, Philipp; Maier, Ezekiel; Saeys, Yvan; Szczurek, Ewa; Myšičková, Alena; Vingron, Martin; Klein, Holger; Kiełbasa, Szymon M.; Knisley, Jeff; Bonnell, Jeff; Knisley, Debra; Kursa, Miron B.; Rudnicki, Witold R.; Bhattacharjee, Madhuchhanda; Sillanpää, Mikko J.; Yeung, James; Meysman, Pieter; Rodríguez, Aminael Sánchez; Engelen, Kristof; Marchal, Kathleen; Huang, Yezhou; Mordelet, Fantine; Hartemink, Alexander; Pinello, Luca; Yuan, Guo-Cheng

2013-01-01

The Gene Promoter Expression Prediction challenge consisted of predicting gene expression from promoter sequences in a previously unknown experimentally generated data set. The challenge was presented to the community in the framework of the sixth Dialogue for Reverse Engineering Assessments and Methods (DREAM6), a community effort to evaluate the status of systems biology modeling methodologies. Nucleotide-specific promoter activity was obtained by measuring fluorescence from promoter sequences fused upstream of a gene for yellow fluorescence protein and inserted in the same genomic site of yeast Saccharomyces cerevisiae. Twenty-one teams submitted results predicting the expression levels of 53 different promoters from yeast ribosomal protein genes. Analysis of participant predictions shows that accurate values for low-expressed and mutated promoters were difficult to obtain, although in the latter case, only when the mutation induced a large change in promoter activity compared to the wild-type sequence. As in previous DREAM challenges, we found that aggregation of participant predictions provided robust results, but did not fare better than the three best algorithms. Finally, this study not only provides a benchmark for the assessment of methods predicting activity of a specific set of promoters from their sequence, but it also shows that the top performing algorithm, which used machine-learning approaches, can be improved by the addition of biological features such as transcription factor binding sites. PMID:23950146

Modern bioanalysis of proteins by electrophoretic techniques.

PubMed

Krizkova, Sona; Ryvolova, Marketa; Masarik, Michal; Zitka, Ondrej; Adam, Vojtech; Hubalek, Jaromir; Eckschlager, Tomas; Kizek, Rene

2014-01-01

In 1957, protein rich in cysteine able to bind cadmium was isolated from horse kidney and named as metallothionein according to its structural properties. Further, this protein and metallothionein-like proteins have been found in tissues of other animal species, yeasts, fungi and plants. MT is as a potential cancer marker in the focus of interest, and its properties, functions, and behavior under various conditions are intensively studied. Our protocol describes separation of two major mammalian isoforms of MT (MT-1 and MT-2) using capillary electrophoresis (CE) coupled with UV detector. This protocol enables separation of MT isoforms and studying of their basic behavior as well as their quantification with detection limit in units of ng per μL. Sodium borate buffer (20 mM, pH 9.5) was optimized as a background electrolyte, and the separation was carried out in fused silica capillary with internal diameter of 75 μm and electric field intensity of 350 V/cm. Optimal detection wavelength was 254 nm.

IE1 and hr facilitate the localization of Bombyx mori nucleopolyhedrovirus ORF8 to specific nuclear sites.

PubMed

Kang, WonKyung; Imai, Noriko; Kawasaki, Yu; Nagamine, Toshihiro; Matsumoto, Shogo

2005-11-01

The Bombyx mori nucleopolyhedrovirus (BmNPV) ORF8 protein has previously been reported to colocalize with IE1 to specific nuclear sites during infection. Transient expression of green fluorescent protein (GFP)-fused ORF8 showed the protein to have cytoplasmic localization, but following BmNPV infection the protein formed foci, suggesting that ORF8 requires some other viral factor(s) for this. Therefore, interacting factors were looked for using the yeast two-hybrid system and IE1 was identified. We mapped the interacting region of ORF8 using a yeast two-hybrid assay. An N-terminal region (residues 1-110) containing a predicted coiled-coil domain interacted with IE1, while a truncated N-terminal region (residues 1-78) that lacks this domain did not. In addition, a protein with a complete deletion of the N-terminal region failed to interact with IE1. These results suggest that the ORF8 N-terminal region containing the coiled-coil domain is required for the interaction with IE1. Next, whether IE1 plays a role in ORF8 localization was investigated. In the presence of IE1, GFP-ORF8 localized to the nucleus. In addition, cotransfection with a plasmid expressing IE1 and a plasmid containing the hr3 element resulted in nuclear foci formation. A GFP-fused ORF8 mutant protein containing the coiled-coil domain, previously shown to interact with IE1, also formed nuclear foci in the presence of IE1 and hr3. However, ORF8 mutant proteins that did not interact with IE1 failed to form nuclear foci. In contrast to wild-type IE1, focus formation was not observed for an IE1 mutant protein that was deficient in hr binding. These results suggest that IE1 and hr facilitate the localization of BmNPV ORF8 to specific nuclear sites.

A Novel Fission Yeast Gene, tht1 +, Is Required for the Fusion of Nuclear Envelopes during Karyogamy

PubMed Central

Tange, Yoshie; Horio, Tetsuya; Shimanuki, Mizuki; Ding, Da-Qiao; Hiraoka, Yasushi; Niwa, Osami

1998-01-01

We have isolated a fission yeast karyogamy mutant, tht1, in which nuclear congression and the association of two spindle pole bodies occurs but the subsequent fusion of nuclear envelopes is blocked. The tht1 mutation does not prevent meiosis, so cells execute meiosis with two unfused nuclei, leading to the production of aberrant asci. The tht1 + gene was cloned and sequenced. Predicted amino acid sequence has no significant homology to previously known proteins but strongly suggests that it is a type I membrane protein. The tht1 + gene is dispensable for vegetative growth and expressed only in conjugating cells. Tht1p is a glycoprotein susceptible to endoglycosilase H digestion. Site- directed mutagenesis showed that the N-glycosylation site, as well as the COOH-terminal region of Tht1p, is essential for its function. A protease protection assay indicated that the COOH terminus is cytoplasmic. Immunocytological analysis using a HA-tagged Tht1p suggested that the protein is localized in nuclear envelopes and in the ER during karyogamy and that its levels are reduced in cells containing fused nuclei. PMID:9442101

Polarized Growth in the Absence of F-Actin in Saccharomyces cerevisiae Exiting Quiescence

PubMed Central

Sahin, Annelise; Daignan-Fornier, Bertrand; Sagot, Isabelle

2008-01-01

Background Polarity establishment and maintenance are crucial for morphogenesis and development. In budding yeast, these two intricate processes involve the superposition of regulatory loops between polarity landmarks, RHO GTPases, actin-mediated vesicles transport and endocytosis. Deciphering the chronology and the significance of each molecular step of polarized growth is therefore very challenging. Principal Findings We have taken advantage of the fact that yeast quiescent cells display actin bodies, a non polarized actin structure, to evaluate the role of F-actin in bud emergence. Here we show that upon exit from quiescence, actin cables are not required for the first steps of polarized growth. We further show that polarized growth can occur in the absence of actin patch-mediated endocytosis. We finally establish, using latrunculin-A, that the first steps of polarized growth do not require any F-actin containing structures. Yet, these structures are required for the formation of a bona fide daughter cell and cell cycle completion. We propose that upon exit from quiescence in the absence of F-actin, secretory vesicles randomly reach the plasma membrane but preferentially dock and fuse where polarity cues are localized, this being sufficient to trigger polarized growth. PMID:18596916

Anti-Amyloidogenic Properties of Some Phenolic Compounds

PubMed Central

Porzoor, Afsaneh; Alford, Benjamin; Hügel, Helmut M.; Grando, Danilla; Caine, Joanne; Macreadie, Ian

2015-01-01

A family of 21 polyphenolic compounds consisting of those found naturally in danshen and their analogues were synthesized and subsequently screened for their anti-amyloidogenic activity against the amyloid beta peptide (Aβ42) of Alzheimer’s disease. After 24 h incubation with Aβ42, five compounds reduced thioflavin T (ThT) fluorescence, indicative of their anti-amyloidogenic propensity (p < 0.001). TEM and immunoblotting analysis also showed that selected compounds were capable of hindering fibril formation even after prolonged incubations. These compounds were also capable of rescuing the yeast cells from toxic changes induced by the chemically synthesized Aβ42. In a second assay, a Saccharomyces cerevisiae AHP1 deletant strain transformed with GFP fused to Aβ42 was treated with these compounds and analyzed by flow cytometry. There was a significant reduction in the green fluorescence intensity associated with 14 compounds. We interpret this result to mean that the compounds had an anti-amyloid-aggregation propensity in the yeast and GFP-Aβ42 was removed by proteolysis. The position and not the number of hydroxyl groups on the aromatic ring was found to be the most important determinant for the anti-amyloidogenic properties. PMID:25898401

AtDMC1, the Arabidopsis homologue of the yeast DMC1 gene: characterization, transposon-induced allelic variation and meiosis-associated expression.

PubMed

Klimyuk, V I; Jones, J D

1997-01-01

Based on homologies between the yeast DMC1 and the lily LIM15 meiosis-specific genes, degenerate PCR primers were designed that amplified the Arabidopsis DMC1 gene (AtDMC1). AtDMC1 genomic DNA (8 kb) was sequenced, and the transcript was characterized by reverse transcriptase-polymerase chain reaction (RT-PCR) and by 5' and 3' RACE (rapid amplification of cDNA ends). The AtDMC1 gene contains 15 exons and 14 introns. RNA in situ hybridization analysis showed that expression of the AtDMC1 is restricted to pollen mother cells in anthers and to megaspore mother cells in ovules. The AtDMC1 promoter was fused to the GUS reporter gene, and conferred meiosis-associated expression in both male and female floral lineages. Comparison of AtDMC1 isolated from Landsberg erecta ecotype to its Columbia allele ArLIM15, revealed the presence of a 1874 bp transposon-like element within the promoter region of ArLIM15. RT-PCR analysis showed that the expression levels of AtDMC1 and ArLIM15 are similar. Possible uses for the AtDMC1 promoter are discussed.

Dicentric breakage at telomere fusions

PubMed Central

Pobiega, Sabrina; Marcand, Stéphane

2010-01-01

Nonhomologous end-joining (NHEJ) inhibition at telomeres ensures that native chromosome ends do not fuse together. But the occurrence and consequences of rare telomere fusions are not well understood. It is notably unclear whether a telomere fusion could be processed to restore telomere ends. Here we address the behavior of individual dicentrics formed by telomere fusion in the yeast Saccharomyces cerevisiae. Our approach was to first stabilize and amplify fusions between two chromosomes by temporarily inactivating one centromere. Next we analyzed dicentric breakage following centromere reactivation. Unexpectedly, dicentrics often break at the telomere fusions during progression through mitosis, a process that restores the parental chromosomes. This unforeseen result suggests a rescue pathway able to process telomere fusions and to back up NHEJ inhibition at telomeres. PMID:20360388

Expression of a plant virus non-structural protein in Saccharomyces cerevisiae causes membrane proliferation and altered mitochondrial morphology.

PubMed

Rubino, L; Di Franco, A; Russo, M

2000-01-01

Carnation Italian ringspot tombusvirus encodes a protein, referred to as 36K, that possesses a mitochondrial targeting signal and two transmembrane segments which are thought to anchor this protein to the outer membrane of the mitochondrial envelope of infected plant cells. To determine the topology of the virus protein inserted in the cell membrane, as well as the sequence requirements for targeting and insertion, an in vivo system was set up in which this could be analysed in the absence of productive virus infection. The 36K protein was expressed in the yeast Saccharomyces cerevisiae in native form or fused to the green fluorescent protein. Using a fluorescence microscope, large green-fluorescing cytoplasmic aggregates were visible which stained red when cells were treated with the vital stain MitoTracker, which is specific for mitochondria. These aggregates were shown by electron microscopy to be composed of either mitochondria or membranes. The latter type was particularly abundant for the construct in which the green fluorescent protein was fused at the N terminus of the 36K protein. Immunoelectron microscopy demonstrated that the viral protein is present in the anomalous aggregates and Western blot analysis of protein extracts showed 36K to be resistant to alkaline, urea or salt extraction, a property of integral membrane proteins.

Genome shuffling of Saccharomyces cerevisiae for enhanced glutathione yield and relative gene expression analysis using fluorescent quantitation reverse transcription polymerase chain reaction.

PubMed

Yin, Hua; Ma, Yanlin; Deng, Yang; Xu, Zhenbo; Liu, Junyan; Zhao, Junfeng; Dong, Jianjun; Yu, Junhong; Chang, Zongming

2016-08-01

Genome shuffling is an efficient and promising approach for the rapid improvement of microbial phenotypes. In this study, genome shuffling was applied to enhance the yield of glutathione produced by Saccharomyces cerevisiae YS86. Six isolates with subtle improvements in glutathione yield were obtained from populations generated by ultraviolet (UV) irradiation and nitrosoguanidine (NTG) mutagenesis. These yeast strains were then subjected to recursive pool-wise protoplast fusion. A strain library that was likely to yield positive colonies was created by fusing the lethal protoplasts obtained from both UV irradiation and heat treatments. After two rounds of genome shuffling, a high-yield recombinant YSF2-19 strain that exhibited 3.2- and 3.3-fold increases in glutathione production in shake flask and fermenter respectively was obtained. Comparative analysis of synthetase gene expression was conducted between the initial and shuffled strains using FQ (fluorescent quantitation) RT-PCR (reverse transcription polymerase chain reaction). Delta CT (threshold cycle) relative quantitation analysis revealed that glutathione synthetase gene (GSH-I) expression at the transcriptional level in the YSF2-19 strain was 9.9-fold greater than in the initial YS86. The shuffled yeast strain has a potential application in brewing, other food, and pharmaceutical industries. Simultaneously, the analysis of improved phenotypes will provide more valuable data for inverse metabolic engineering. Copyright © 2016 Elsevier B.V. All rights reserved.

Functional analysis of AoAtg11 in selective autophagy in the filamentous fungus Aspergillus oryzae.

PubMed

Tadokoro, Takayuki; Kikuma, Takashi; Kitamoto, Katsuhiko

2015-07-01

Autophagy is a highly conserved cellular degradation process in eukaryotes and consists of both non-selective and selective types. Selective autophagic processes include pexophagy, mitophagy, and the cytoplasm-to-vacuole targeting (Cvt) pathway of yeast, in which particular vacuolar proteins, such as aminopeptidase I (Ape1), are selectively transported to vacuoles. Although selective autophagy has been mainly studied in the yeasts Saccharomyces cerevisiae and Pichia pastoris, there is evidence for selective autophagy in filamentous fungi; however, the details are poorly understood. In S. cerevisiae, Atg11 is a selective autophagy-specific protein that recognizes and transports substrates to the pre-autophagosomal structure (PAS). Here, we first identified an ATG11 homologue in the filamentous fungus Aspergillus oryzae and analyzed the localization of the corresponding protein, designated AoAtg11, fused to enhanced green fluorescent protein (EGFP). Imaging analysis revealed that AoAtg11-EGFP was localized to PAS-like structures. We next constructed an Aoatg11 disruptant of A. oryzae and showed that AoAtg11 is involved in pexophagy and mitophagy. In addition, AoAtg11 was found to be dispensable for non-selective autophagy and for transporting AoApe1 to vacuoles. Taken together, these results suggest that AoAtg11 is a selective autophagy-specific protein in A. oryzae, and has distinct molecular functions from that of S. cerevisiae Atg11. Copyright © 2015 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

Activation of Rab GTPase Sec4 by its GEF Sec2 is required for prospore membrane formation during sporulation in yeast Saccharomyces cerevisiae.

PubMed

Suda, Yasuyuki; Tachikawa, Hiroyuki; Inoue, Ichiro; Kurita, Tomokazu; Saito, Chieko; Kurokawa, Kazuo; Nakano, Akihiko; Irie, Kenji

2018-02-01

Sec2 activates Sec4 Rab GTPase as a guanine nucleotide exchange factor for the recruitment of downstream effectors to facilitate tethering and fusion of post-Golgi vesicles at the plasma membrane. During the meiosis and sporulation of budding yeast, post-Golgi vesicles are transported to and fused at the spindle pole body (SPB) to form a de novo membrane, called the prospore membrane. Previous studies have revealed the role of the SPB outer surface called the meiotic outer plaque (MOP) in docking and fusion of post-Golgi vesicles. However, the upstream molecular machinery for post-Golgi vesicular fusion that facilitates prospore membrane formation remains enigmatic. Here, we demonstrate that the GTP exchange factor for Sec4, Sec2, participates in the formation of the prospore membrane. A conditional mutant in which the SEC2 expression is shut off during sporulation showed sporulation defects. Inactivation of Sec2 caused Sec4 targeting defects along the prospore membranes, thereby causing insufficient targeting of downstream effectors and cargo proteins to the prospore membrane. These results suggest that the activation of Sec4 by Sec2 is required for the efficient supply of post-Golgi vesicles to the prospore membrane and thus for prospore membrane formation/extension and subsequent deposition of spore wall materials. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

A visual screen for diet-regulated proteins in the Drosophila ovary using GFP protein trap lines.

PubMed

Hsu, Hwei-Jan; Drummond-Barbosa, Daniela

2017-01-01

The effect of diet on reproduction is well documented in a large number of organisms; however, much remains to be learned about the molecular mechanisms underlying this connection. The Drosophila ovary has a well described, fast and largely reversible response to diet. Ovarian stem cells and their progeny proliferate and grow faster on a yeast-rich diet than on a yeast-free (poor) diet, and death of early germline cysts, degeneration of early vitellogenic follicles and partial block in ovulation further contribute to the ∼60-fold decrease in egg laying observed on a poor diet. Multiple diet-dependent factors, including insulin-like peptides, the steroid ecdysone, the nutrient sensor Target of Rapamycin, AMP-dependent kinase, and adipocyte factors mediate this complex response. Here, we describe the results of a visual screen using a collection of green fluorescent protein (GFP) protein trap lines to identify additional factors potentially involved in this response. In each GFP protein trap line, an artificial GFP exon is fused in frame to an endogenous protein, such that the GFP fusion pattern parallels the levels and subcellular localization of the corresponding native protein. We identified 53 GFP-tagged proteins that exhibit changes in levels and/or subcellular localization in the ovary at 12-16 hours after switching females from rich to poor diets, suggesting them as potential candidates for future functional studies. Copyright © 2017 Elsevier B.V. All rights reserved.

Usefulness of 2D fusion of postmortem CT and antemortem chest radiography studies for human identification.

PubMed

Shinkawa, Norihiro; Hirai, Toshinori; Nishii, Ryuichi; Yukawa, Nobuhiro

2017-06-01

To determine the feasibility of human identification through the two-dimensional (2D) fusion of postmortem computed tomography (PMCT) and antemortem chest radiography. The study population consisted of 15 subjects who had undergone chest radiography studies more than 12 months before death. Fused images in which a chest radiograph was fused with a PMCT image were obtained for those subjects using a workstation, and the minimum distance gaps between corresponding anatomical landmarks (located at soft tissue and bone sites) in the images obtained with the two modalities were calculated. For each fused image, the mean of all these minimum distance gaps was recorded as the mean distance gap (MDG). For each subject, the MDG obtained for the same-subject fused image (i.e., where both of the images that were fused derived from that subject) was compared with the MDGs for different-subject fused images (i.e., where only one of the images that were fused derived from that subject; the other image derived from a different subject) in order to determine whether same-subject fused images can be reliably distinguished from different-subject fused images. The MDGs of the same-subject fused images were found to be significantly smaller than the MDGs of the different-subject fused images (p < 0.01). When bone landmarks were used, the same-subject fused image was found to be the fused image with the lowest MDG for 33.3% of the subjects, the fused image with the lowest or second-lowest MDG for 73.3% of the subjects, and the fused image with the lowest, second-lowest, or third-lowest MDG for 86.7% of the subjects. The application of bone landmarks rather than soft-tissue landmarks made it significantly more likely that, for each subject, the same-subject fused image would have the lowest MDG (or one of the lowest MDGs) of all the fused images compared (p < 0.05). The 2D fusion of antemortem chest radiography and postmortem CT images may assist in human identification.

Chromatibody, a novel non-invasive molecular tool to explore and manipulate chromatin in living cells

PubMed Central

Jullien, Denis; Vignard, Julien; Fedor, Yoann; Béry, Nicolas; Olichon, Aurélien; Crozatier, Michèle; Erard, Monique; Cassard, Hervé; Ducommun, Bernard; Salles, Bernard

2016-01-01

ABSTRACT Chromatin function is involved in many cellular processes, its visualization or modification being essential in many developmental or cellular studies. Here, we present the characterization of chromatibody, a chromatin-binding single-domain, and explore its use in living cells. This non-intercalating tool specifically binds the heterodimer of H2A–H2B histones and displays a versatile reactivity, specifically labeling chromatin from yeast to mammals. We show that this genetically encoded probe, when fused to fluorescent proteins, allows non-invasive real-time chromatin imaging. Chromatibody is a dynamic chromatin probe that can be modulated. Finally, chromatibody is an efficient tool to target an enzymatic activity to the nucleosome, such as the DNA damage-dependent H2A ubiquitylation, which can modify this epigenetic mark at the scale of the genome and result in DNA damage signaling and repair defects. Taken together, these results identify chromatibody as a universal non-invasive tool for either in vivo chromatin imaging or to manipulate the chromatin landscape. PMID:27206857

Mitochondrial recombination increases with age in Podospora anserina.

PubMed

van Diepeningen, Anne D; Goedbloed, Daniël J; Slakhorst, S Marijke; Koopmanschap, A Bertha; Maas, Marc F P M; Hoekstra, Rolf F; Debets, Alfons J M

2010-05-01

With uniparental inheritance of mitochondria, there seems little reason for homologous recombination in mitochondria, but the machinery for mitochondrial recombination is quite well-conserved in many eukaryote species. In fungi and yeasts heteroplasmons may be formed when strains fuse and transfer of organelles takes place, making it possible to study mitochondrial recombination when introduced mitochondria contain different markers. A survey of wild-type isolates from a local population of the filamentous fungus Podospora anserina for the presence of seven optional mitochondrial introns indicated that mitochondrial recombination does take place in nature. Moreover the recombination frequency appeared to be correlated with age: the more rapidly ageing fraction of the population had a significantly lower linkage disequilibrium indicating more recombination. Direct confrontation experiments with heterokaryon incompatible strains with different mitochondrial markers at different (relative) age confirmed that mitochondrial recombination increases with age. We propose that with increasing mitochondrial damage over time, mitochondrial recombination - even within a homoplasmic population of mitochondria - is a mechanism that may restore mitochondrial function. Copyright 2010 Elsevier Ireland Ltd. All rights reserved.

Citrobacter amalonaticus Phytase on the Cell Surface of Pichia pastoris Exhibits High pH Stability as a Promising Potential Feed Supplement

PubMed Central

Li, Cheng; Lin, Ying; Huang, Yuanyuan; Liu, Xiaoxiao; Liang, Shuli

2014-01-01

Phytase expressed and anchored on the cell surface of Pichia pastoris avoids the expensive and time-consuming steps of protein purification and separation. Furthermore, yeast cells with anchored phytase can be used as a whole-cell biocatalyst. In this study, the phytase gene of Citrobacter amalonaticus was fused with the Pichia pastoris glycosylphosphatidylinositol (GPI)-anchored glycoprotein homologue GCW61. Phytase exposed on the cell surface exhibits a high activity of 6413.5 U/g, with an optimal temperature of 60°C. In contrast to secreted phytase, which has an optimal pH of 5.0, phytase presented on the cell surface is characterized by an optimal pH of 3.0. Moreover, our data demonstrate that phytase anchored on the cell surface exhibits higher pH stability than its secreted counterpart. Interestingly, our in vitro digestion experiments demonstrate that phytase attached to the cell surface is a more efficient enzyme than secreted phytase. PMID:25490768

Citrobacter amalonaticus phytase on the cell surface of Pichia pastoris exhibits high pH stability as a promising potential feed supplement.

PubMed

Li, Cheng; Lin, Ying; Huang, Yuanyuan; Liu, Xiaoxiao; Liang, Shuli

2014-01-01

Phytase expressed and anchored on the cell surface of Pichia pastoris avoids the expensive and time-consuming steps of protein purification and separation. Furthermore, yeast cells with anchored phytase can be used as a whole-cell biocatalyst. In this study, the phytase gene of Citrobacter amalonaticus was fused with the Pichia pastoris glycosylphosphatidylinositol (GPI)-anchored glycoprotein homologue GCW61. Phytase exposed on the cell surface exhibits a high activity of 6413.5 U/g, with an optimal temperature of 60°C. In contrast to secreted phytase, which has an optimal pH of 5.0, phytase presented on the cell surface is characterized by an optimal pH of 3.0. Moreover, our data demonstrate that phytase anchored on the cell surface exhibits higher pH stability than its secreted counterpart. Interestingly, our in vitro digestion experiments demonstrate that phytase attached to the cell surface is a more efficient enzyme than secreted phytase.

Exploring the transcription activator-like effectors scaffold versatility to expand the toolbox of designer nucleases

PubMed Central

2014-01-01

Background The past decade has seen the emergence of several molecular tools that render possible modification of cellular functions through accurate and easy addition, removal, or exchange of genomic DNA sequences. Among these technologies, transcription activator-like effectors (TALE) has turned out to be one of the most versatile and incredibly robust platform for generating targeted molecular tools as demonstrated by fusion to various domains such as transcription activator, repressor and nucleases. Results In this study, we generated a novel nuclease architecture based on the transcription activator-like effector scaffold. In contrast to the existing Tail to Tail (TtT) and head to Head (HtH) nuclease architectures based on the symmetrical association of two TALE DNA binding domains fused to the C-terminal (TtT) or N-terminal (HtH) end of FokI, this novel architecture consists of the asymmetrical association of two different engineered TALE DNA binding domains fused to the N- and C-terminal ends of FokI (TALE::FokI and FokI::TALE scaffolds respectively). The characterization of this novel Tail to Head (TtH) architecture in yeast enabled us to demonstrate its nuclease activity and define its optimal target configuration. We further showed that this architecture was able to promote substantial level of targeted mutagenesis at three endogenous loci present in two different mammalian cell lines. Conclusion Our results demonstrated that this novel functional TtH architecture which requires binding to only one DNA strand of a given endogenous locus has the potential to extend the targeting possibility of FokI-based TALE nucleases. PMID:24997498

T-DNA transfer and integration in the ectomycorrhizal fungus Suillus bovinus using hygromycin B as a selectable marker.

PubMed

Hanif, Mubashir; Pardo, Alejandro Guillermo; Gorfer, Markus; Raudaskoski, Marjatta

2002-06-01

The T-DNA of Agrobacterium tumefaciens can be transferred to plants, yeasts, fungi and human cells. Using this system, dikaryotic mycelium of the ectomycorrhizal fungus Suillus bovinus was transformed with recombinant hygromycin B phosphotransferase (hph)and enhanced green fluorescent protein (EGFP) genes fused with a heterologous fungal promoter and CaMV35S terminator. Transformation resulted in hygromycin B-resistant clones, which were mitotically stable. Putative transformants were analysed for the presence of hph and EGFP genes by PCR and Southern analysis. The latter analysis proved both multiple- and single-copy integrations of the genes in the S. bovinus genome. A. tumeficiens transformation should make possible the development of tagged mutagenesis and targeted gene disruption technology for S. bovinus.

Extracellular expression of YlLip11 with a native signal peptide from Yarrowia lipolytica MSR80 in three different yeast hosts.

PubMed

Kumari, Arti; Baronian, Keith; Kunze, Gotthard; Gupta, Rani

2015-06-01

Lipase YlLip11 from Yarrowia lipolytica was expressed with a signal peptide encoding sequence in Arxula adeninivorans, Saccharomyces cerevisiae and Hansenula polymorpha using the Xplor®2 transformation/expression platform and an expression module with the constitutive Arxula-derived TEF1 promoter. The YlLip11 signal peptide was functional in all of the yeast hosts with 97% of the recombinant enzyme being secreted into the culture medium. However, recombinant YlLip11 with His Tag fused at C-terminal was not active. The best recombinant YlLip11 producing A. adeninivorans G1212/YRC102-YlLip11 transformant cultivated in shake flasks produced 2654 U/L lipase, followed by S. cerevisiae SEY6210/YRC103-YlLip11 (1632U/L) and H. polymorpha RB11/YRC103-YlLip11 (1144U/L). Although the biochemical parameters of YlLip11 synthesized in different hosts were similar, their glycosylation level and thermo stability differed. The protein synthesized by the H. polymorpha transformant had the highest degree of glycosylation and with a t1/2 of 60min at 70°C, exhibited the highest thermostability. Copyright © 2015 Elsevier Inc. All rights reserved.

Fundamental Use of Surgical Energy (FUSE) certification: validation and predictors of success.

PubMed

Robinson, Thomas N; Olasky, Jaisa; Young, Patricia; Feldman, Liane S; Fuchshuber, Pascal R; Jones, Stephanie B; Madani, Amin; Brunt, Michael; Mikami, Dean; Jackson, Gretchen P; Mischna, Jessica; Schwaitzberg, Steven; Jones, Daniel B

2016-03-01

The Fundamental Use of Surgical Energy (FUSE) program includes a Web-based didactic curriculum and a high-stakes multiple-choice question examination with the goal to provide certification of knowledge on the safe use of surgical energy-based devices. The purpose of this study was (1) to set a passing score through a psychometrically sound process and (2) to determine what pretest factors predicted passing the FUSE examination. Beta-testing of multiple-choice questions on 62 topics of importance to the safe use of surgical energy-based devices was performed. Eligible test takers were physicians with a minimum of 1 year of surgical training who were recruited by FUSE task force members. A pretest survey collected baseline information. A total of 227 individuals completed the FUSE beta-test, and 208 completed the pretest survey. The passing/cut score for the first test form of the FUSE multiple-choice examination was determined using the modified Angoff methodology and for the second test form was determined using a linear equating methodology. The overall passing rate across the two examination forms was 81.5%. Self-reported time studying the FUSE Web-based curriculum for a minimum of >2 h was associated with a passing examination score (p < 0.001). Performance was not different based on increased years of surgical practice (p = 0.363), self-reported expertise on one or more types of energy-based devices (p = 0.683), participation in the FUSE postgraduate course (p = 0.426), or having reviewed the FUSE manual (p = 0.428). Logistic regression found that studying the FUSE didactics for >2 h predicted a passing score (OR 3.61; 95% CI 1.44-9.05; p = 0.006) independent of the other baseline characteristics recorded. The development of the FUSE examination, including the passing score, followed a psychometrically sound process. Self-reported time studying the FUSE curriculum predicted a passing score independent of other pretest characteristics such as years in practice and self-reported expertise.

Actin cable dynamics in budding yeast

PubMed Central

Yang, Hyeong-Cheol; Pon, Liza A.

2002-01-01

Actin cables, bundles of actin filaments that align along the long axis of budding yeast, are crucial for establishment of cell polarity. We fused green fluorescent protein (GFP) to actin binding protein 140 (Abp140p) and visualized actin cable dynamics in living yeast. We detected two populations of actin cables: (i) bud-associated cables, which extend from the bud along the mother-bud axis, and (ii) randomly oriented cables, which are relatively short. Time-lapse imaging of Abp140p–GFP revealed an apparent increase in the length of bud-associated actin cables. Analysis of movement of Abp140p–GFP fiduciary marks on bud-associated cables and fluorescence loss in photobleaching experiments revealed that this apparent elongation occurs by assembly of new material at the end of the cable within the bud and movement of the opposite end of the cable toward the tip of the mother cell distal to the bud. The rate of extension of the tip of an elongating actin cable is 0.29 ± 0.08 μm/s. Latrunculin A (Lat-A) treatment completely blocked this process. We also observed movement of randomly oriented cables around the cortex of cells at a rate of 0.59 ± 0.14 μm/s. Mild treatment with Lat-A did not affect the velocity of movement of randomly oriented cables. However, Lat-A treatment did increase the number of randomly oriented, motile cables per cell. Our observations suggest that establishment of bud-associated actin cables during the cell cycle is accomplished not by realignment of existing cables but by assembly of new cables within the bud or bud neck, followed by elongation. PMID:11805329

Unraveling the function of Arabidopsis thaliana OS9 in the endoplasmic reticulum-associated degradation of glycoproteins.

PubMed

Hüttner, Silvia; Veit, Christiane; Schoberer, Jennifer; Grass, Josephine; Strasser, Richard

2012-05-01

In the endoplasmic reticulum, immature polypeptides coincide with terminally misfolded proteins. Consequently, cells need a well-balanced quality control system, which decides about the fate of individual proteins and maintains protein homeostasis. Misfolded and unassembled proteins are sent for destruction via the endoplasmic reticulum-associated degradation (ERAD) machinery to prevent the accumulation of potentially toxic protein aggregates. Here, we report the identification of Arabidopsis thaliana OS9 as a component of the plant ERAD pathway. OS9 is an ER-resident glycoprotein containing a mannose-6-phosphate receptor homology domain, which is also found in yeast and mammalian lectins involved in ERAD. OS9 fused to the C-terminal domain of YOS9 can complement the ERAD defect of the corresponding yeast Δyos9 mutant. An A. thaliana OS9 loss-of-function line suppresses the severe growth phenotype of the bri1-5 and bri1-9 mutant plants, which harbour mutated forms of the brassinosteroid receptor BRI1. Co-immunoprecipitation studies demonstrated that OS9 associates with Arabidopsis SEL1L/HRD3, which is part of the plant ERAD complex and with the ERAD substrates BRI1-5 and BRI1-9, but only the binding to BRI1-5 occurs in a glycan-dependent way. OS9-deficiency results in activation of the unfolded protein response and reduces salt tolerance, highlighting the role of OS9 during ER stress. We propose that OS9 is a component of the plant ERAD machinery and may act specifically in the glycoprotein degradation pathway.

Identification of new members of Fertilisation Independent Seed Polycomb Group pathway involved in the control of seed development in Arabidopsis thaliana.

PubMed

Guitton, Anne-Elisabeth; Page, Damian R; Chambrier, Pierre; Lionnet, Claire; Faure, Jean-Emmanuel; Grossniklaus, Ueli; Berger, Frédéric

2004-06-01

In higher plants, double fertilisation initiates seed development. One sperm cell fuses with the egg cell and gives rise to the embryo, the second sperm cell fuses with the central cell and gives rise to the endosperm. The endosperm develops as a syncytium with the gradual organisation of domains along an anteroposterior axis defined by the position of the embryo at the anterior pole and by the attachment to the placenta at the posterior pole. We report that ontogenesis of the posterior pole in Arabidopsis thaliana involves oriented migration of nuclei in the syncytium. We show that this migration is impaired in mutants of the three founding members of the FERTILIZATION INDEPENDENT SEED (FIS) class, MEDEA (MEA), FIS2 and FERTILIZATION INDEPENDENT ENDOSPERM (FIE). A screen based on a green fluorescent protein (GFP) reporter line allowed us to identify two new loci in the FIS pathway, medicis and borgia. We have cloned the MEDICIS gene and show that it encodes the Arabidopsis homologue of the yeast WD40 domain protein MULTICOPY SUPRESSOR OF IRA (MSI1). The mutations at the new fis loci cause the same cellular defects in endosperm development as other fis mutations, including parthenogenetic development, absence of cellularisation, ectopic development of posterior structures and overexpression of the GFP marker.

Evaluation of sterol transport from the endoplasmic reticulum to mitochondria using mitochondrially targeted bacterial sterol acyltransferase in Saccharomyces cerevisiae.

PubMed

Tian, Siqi; Ohta, Akinori; Horiuchi, Hiroyuki; Fukuda, Ryouichi

2015-01-01

To elucidate the mechanism of interorganelle sterol transport, a system to evaluate sterol transport from the endoplasmic reticulum (ER) to the mitochondria was constructed. A bacterial glycerophospholipid: cholesterol acyltransferase fused with a mitochondria-targeting sequence and a membrane-spanning domain of the mitochondrial inner membrane protein Pet100 and enhanced green fluorescent protein was expressed in a Saccharomyces cerevisiae mutant deleted for ARE1 and ARE2 encoding acyl-CoA:sterol acyltransferases. Microscopic observation and subcellular fractionation suggested that this fusion protein, which was named mito-SatA-EGFP, was localized in the mitochondria. Steryl esters were synthesized in the mutant expressing mito-SatA-EGFP. This system will be applicable for evaluations of sterol transport from the ER to the mitochondria in yeast by examining sterol esterification in the mitochondria.

Fiber Fabry-Perot Interferometric Sensor for the Measurement of Electric Current Flowing through a Fuse

NASA Astrophysics Data System (ADS)

Park, Jaehee

2007-06-01

A fiber Fabry-Perot inteferometric sensor bonded close to a fusing element has been studied for the measurement of electric current flowing through a fuse. The phase shift of the sensor output signal is proportional to the square of the electric current passing through the fuse and the sensitivity is 0.827°/mA2.

Yeast Surface Display of Two Proteins Previously Shown to Be Protective Against White Spot Syndrome Virus (WSSV) in Shrimp.

PubMed

Ananphongmanee, Vorawit; Srisala, Jiraporn; Sritunyalucksana, Kallaya; Boonchird, Chuenchit

2015-01-01

Cell surface display using the yeasts Saccharomyces cerevisiae and Pichia pastoris has been extensively developed for application in bioindustrial processes. Due to the rigid structure of their cell walls, a number of proteins have been successfully displayed on their cell surfaces. It was previously reported that the viral binding protein Rab7 from the giant tiger shrimp Penaeus monodon (PmRab7) and its binding partner envelope protein VP28 of white spot syndrome virus (WSSV) could independently protect shrimp against WSSV infection. Thus, we aimed to display these two proteins independently on the cell surfaces of 2 yeast clones with the ultimate goal of using a mixture of the two clones as an orally deliverable, antiviral agent to protect shrimp against WSSV infection. PmRab7 and VP28 were modified by N-terminal tagging to the C-terminal half of S. cerevisiae α-agglutinin. DNA fragments, harboring fused-gene expression cassettes under control of an alcohol oxidase I (AOX1) promoter were constructed and used to transform the yeast cells. Immunofluorescence microscopy with antibodies specific to both proteins demonstrated that mutated PmRab7 (mPmRab7) and partial VP28 (pVP28) were localized on the cell surfaces of the respective clones, and fluorescence intensity for each was significantly higher than that of control cells by flow cytometry. Enzyme-linked immunosorbant assay (ELISA) using cells displaying mPmRab7 or pVP28 revealed that the binding of specific antibodies for each was dose-dependent, and could be saturated. In addition, the binding of mPmRab7-expressing cells with free VP28, and vice versa was dose dependent. Binding between the two surface-expressed proteins was confirmed by an assay showing agglutination between cells expressing complementary mPmRab7 and pVP28. In summary, our genetically engineered P. pastoris can display biologically active mPmRab7 and pVP28 and is now ready for evaluation of efficacy in protecting shrimp against WSSV by oral administration.

Rho2 Palmitoylation Is Required for Plasma Membrane Localization and Proper Signaling to the Fission Yeast Cell Integrity Mitogen-Activated Protein Kinase Pathway

PubMed Central

Sánchez-Mir, Laura; Franco, Alejandro; Martín-García, Rebeca; Madrid, Marisa; Vicente-Soler, Jero; Soto, Teresa; Gacto, Mariano; Pérez, Pilar

2014-01-01

The fission yeast small GTPase Rho2 regulates morphogenesis and is an upstream activator of the cell integrity pathway, whose key element, mitogen-activated protein kinase (MAPK) Pmk1, becomes activated by multiple environmental stimuli and controls several cellular functions. Here we demonstrate that farnesylated Rho2 becomes palmitoylated in vivo at cysteine-196 within its carboxyl end and that this modification allows its specific targeting to the plasma membrane. Unlike that of other palmitoylated and prenylated GTPases, the Rho2 control of morphogenesis and Pmk1 activity is strictly dependent upon plasma membrane localization and is not found in other cellular membranes. Indeed, artificial plasma membrane targeting bypassed the Rho2 need for palmitoylation in order to signal. Detailed functional analysis of Rho2 chimeras fused to the carboxyl end from the essential GTPase Rho1 showed that GTPase palmitoylation is partially dependent on the prenylation context and confirmed that Rho2 signaling is independent of Rho GTP dissociation inhibitor (GDI) function. We further demonstrate that Rho2 is an in vivo substrate for DHHC family acyltransferase Erf2 palmitoyltransferase. Remarkably, Rho3, another Erf2 target, negatively regulates Pmk1 activity in a Rho2-independent fashion, thus revealing the existence of cross talk whereby both GTPases antagonistically modulate the activity of this MAPK cascade. PMID:24820419

Fiber fuse behavior in kW-level continuous-wave double-clad field laser

NASA Astrophysics Data System (ADS)

Jun-Yi, Sun; Qi-Rong, Xiao; Dan, Li; Xue-Jiao, Wang; Hai-Tao, Zhang; Ma-Li, Gong; Ping, Yan

2016-01-01

In this study, original experimental data for fiber fuse in kW-level continuous-wave (CW) high power double-clad fiber (DCF) laser are reported. The propagating velocity of the fuse is 9.68 m/s in a 3.1-kW Yb-doped DCF laser. Three other cases in Yb-doped DCF are also observed. We think that the ignition of fiber fuse is caused by thermal mechanism, and the formation of bullet-shaped tracks is attributed to the optical discharge and temperature gradient. The inducements of initial fuse and formation of bullet-shaped voids are analyzed. This investigation of fiber fuse helps better understand the fiber fuse behavior, in order to avoid the catastrophic destruction caused by fiber fuse in high power fiber laser. Project supported by the Key Laboratory of Science and Technology on High Energy Laser and China Academy of Engineering Physics (Grant No. 2014HEL02) and the National Natural Science Foundation of China (Grant No. 61307057).

Yarrowia lipolytica vesicle-mediated protein transport pathways

PubMed Central

Swennen, Dominique; Beckerich, Jean-Marie

2007-01-01

Background Protein secretion is a universal cellular process involving vesicles which bud and fuse between organelles to bring proteins to their final destination. Vesicle budding is mediated by protein coats; vesicle targeting and fusion depend on Rab GTPase, tethering factors and SNARE complexes. The Génolevures II sequencing project made available entire genome sequences of four hemiascomycetous yeasts, Yarrowia lipolytica, Debaryomyces hansenii, Kluyveromyces lactis and Candida glabrata. Y. lipolytica is a dimorphic yeast and has good capacities to secrete proteins. The translocation of nascent protein through the endoplasmic reticulum membrane was well studied in Y. lipolytica and is largely co-translational as in the mammalian protein secretion pathway. Results We identified S. cerevisiae proteins involved in vesicular secretion and these protein sequences were used for the BLAST searches against Génolevures protein database (Y. lipolytica, C. glabrata, K. lactis and D. hansenii). These proteins are well conserved between these yeasts and Saccharomyces cerevisiae. We note several specificities of Y. lipolytica which may be related to its good protein secretion capacities and to its dimorphic aspect. An expansion of the Y. lipolytica Rab protein family was observed with autoBLAST and the Rab2- and Rab4-related members were identified with BLAST against NCBI protein database. An expansion of this family is also found in filamentous fungi and may reflect the greater complexity of the Y. lipolytica secretion pathway. The Rab4p-related protein may play a role in membrane recycling as rab4 deleted strain shows a modification of colony morphology, dimorphic transition and permeability. Similarly, we find three copies of the gene (SSO) encoding the plasma membrane SNARE protein. Quantification of the percentages of proteins with the greatest homology between S. cerevisiae, Y. lipolytica and animal homologues involved in vesicular transport shows that 40% of Y. lipolytica proteins are closer to animal ones, whereas they are only 13% in the case of S. cerevisiae. Conclusion These results provide further support for the idea, previously noted about the endoplasmic reticulum translocation pathway, that Y. lipolytica is more representative of vesicular secretion of animals and other fungi than is S. cerevisiae. PMID:17997821

The structure of transcription termination factor Nrd1 reveals an original mode for GUAA recognition

PubMed Central

Franco-Echevarría, Elsa; González-Polo, Noelia; Zorrilla, Silvia; Martínez-Lumbreras, Santiago; Santiveri, Clara M.; Campos-Olivas, Ramón; Sánchez, Mar; Calvo, Olga

2017-01-01

Abstract Transcription termination of non-coding RNAs is regulated in yeast by a complex of three RNA binding proteins: Nrd1, Nab3 and Sen1. Nrd1 is central in this process by interacting with Rbp1 of RNA polymerase II, Trf4 of TRAMP and GUAA/G terminator sequences. We lack structural data for the last of these binding events. We determined the structures of Nrd1 RNA binding domain and its complexes with three GUAA-containing RNAs, characterized RNA binding energetics and tested rationally designed mutants in vivo. The Nrd1 structure shows an RRM domain fused with a second α/β domain that we name split domain (SD), because it is formed by two non-consecutive segments at each side of the RRM. The GUAA interacts with both domains and with a pocket of water molecules, trapped between the two stacking adenines and the SD. Comprehensive binding studies demonstrate for the first time that Nrd1 has a slight preference for GUAA over GUAG and genetic and functional studies suggest that Nrd1 RNA binding domain might play further roles in non-coding RNAs transcription termination. PMID:28973465

Yeast flocculation: New story in fuel ethanol production.

PubMed

Zhao, X Q; Bai, F W

2009-01-01

Yeast flocculation has been used in the brewing industry to facilitate biomass recovery for a long time, and thus its mechanism of yeast flocculation has been intensively studied. However, the application of flocculating yeast in ethanol production garnered attention mainly in the 1980s and 1990s. In this article, updated research progress in the molecular mechanism of yeast flocculation and the impact of environmental conditions on yeast flocculation are reviewed. Construction of flocculating yeast strains by genetic approach and utilization of yeast flocculation for ethanol production from various feedstocks were presented. The concept of self-immobilized yeast cells through their flocculation is revisited through a case study of continuous ethanol fermentation with the flocculating yeast SPSC01, and their technical and economic advantages are highlighted by comparing with yeast cells immobilized with supporting materials and regular free yeast cells as well. Taking the flocculating yeast SPSC01 as an example, the ethanol tolerance of the flocculating yeast was also discussed.

A plasmid collection for PCR-based gene targeting in the filamentous ascomycete Ashbya gossypii.

PubMed

Kaufmann, Andreas

2009-08-01

PCR-based gene targeting with heterologous markers is an efficient method to delete genes, generate gene fusions, and modulate gene expression. For the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, several plasmid collections are available covering a wide range of tags and markers. For several reasons, many of these cassettes cannot be used in the filamentous ascomycete Ashbya gossypii. This article describes the construction of 93 heterologous modules for C- and N-terminal tagging and promoter replacements in A. gossypii. The performance of 12 different fluorescent tags was evaluated by monitoring their brightness, detectability, and photostability when fused to the myosin light-chain protein Mlc2. Furthermore, the thiamine-repressible S. cerevisiae THI13 promoter was established to regulate gene expression in A. gossypii. This collection will help accelerate analysis of gene function in A. gossypii and in other ascomycetes where S. cerevisiae promoter elements are functional.

pHlash: A New Genetically Encoded and Ratiometric Luminescence Sensor of Intracellular pH

PubMed Central

Robertson, J. Brian; Johnson, Carl Hirschie

2012-01-01

We report the development of a genetically encodable and ratiometic pH probe named “pHlash” that utilizes Bioluminescence Resonance Energy Transfer (BRET) rather than fluorescence excitation. The pHlash sensor–composed of a donor luciferase that is genetically fused to a Venus fluorophore–exhibits pH dependence of its spectral emission in vitro. When expressed in either yeast or mammalian cells, pHlash reports basal pH and cytosolic acidification in vivo. Its spectral ratio response is H+ specific; neither Ca++, Mg++, Na+, nor K+ changes the spectral form of its luminescence emission. Moreover, it can be used to image pH in single cells. This is the first BRET-based sensor of H+ ions, and it should allow the approximation of pH in cytosolic and organellar compartments in applications where current pH probes are inadequate. PMID:22905204

pHlash: a new genetically encoded and ratiometric luminescence sensor of intracellular pH.

PubMed

Zhang, Yunfei; Xie, Qiguang; Robertson, J Brian; Johnson, Carl Hirschie

2012-01-01

We report the development of a genetically encodable and ratiometic pH probe named "pHlash" that utilizes Bioluminescence Resonance Energy Transfer (BRET) rather than fluorescence excitation. The pHlash sensor-composed of a donor luciferase that is genetically fused to a Venus fluorophore-exhibits pH dependence of its spectral emission in vitro. When expressed in either yeast or mammalian cells, pHlash reports basal pH and cytosolic acidification in vivo. Its spectral ratio response is H(+) specific; neither Ca(++), Mg(++), Na(+), nor K(+) changes the spectral form of its luminescence emission. Moreover, it can be used to image pH in single cells. This is the first BRET-based sensor of H(+) ions, and it should allow the approximation of pH in cytosolic and organellar compartments in applications where current pH probes are inadequate.

The Effect of α-Mating Factor Secretion Signal Mutations on Recombinant Protein Expression in Pichia pastoris

PubMed Central

Lin-Cereghino, Geoff P.; Stark, Carolyn M.; Kim, Daniel; Chang, Jennifer; Shaheen, Nadia; Poerwanto, Hansel; Agari, Kimiko; Moua, Pachai; Low, Lauren K.; Tran, Namphuong; Huang, Amy D.; Nattestad, Maria; Oshiro, Kristin T.; Chang, John William; Chavan, Archana; Tsai, Jerry W.; Lin-Cereghino, Joan

2013-01-01

The methylotrophic yeast, Pichia pastoris, has been genetically engineered to produce many heterologous proteins for industrial and research purposes. In order to secrete proteins for easier purification from the extracellular medium, the coding sequence of recombinant proteins are initially fused to the Saccharomyces cerevisiae α-mating factor secretion signal leader. Extensive site-directed mutagenesis of the prepro region of the α-mating factor secretion signal sequence was performed in order to determine the effects of various deletions and substitutions on expression. Though some mutations clearly dampened protein expression, deletion of amino acids 57-70, corresponding to the predicted 3rd alpha helix of α-mating factor secretion signal, increased secretion of reporter proteins horseradish peroxidase and lipase at least 50% in small-scale cultures. These findings raise the possibility that the secretory efficiency of the leader can be further enhanced in the future. PMID:23454485

The research progress of large-aperture fused silica for high power laser

NASA Astrophysics Data System (ADS)

Shao, Zhufeng; Wang, Yufen; Xiang, Zaikui; Rao, Chuandong

2016-03-01

Because of its excellent optical performance, the fused silica is widely used in laser industry. In addition, the fused silica can withstand high power laser, due to its pure component, and the performance is most outstanding within all types of glasses. So fused silica can be used for optical lens in high power laser field. From the manufacturing process stand point, the fused silica can be categorized to four types: type Ⅰ, type Ⅱ, type Ⅲ, and type Ⅳ. The fused silica of type Ⅰand type Ⅱ is made through melting silica sand in graphite furnace or oxyhydrogen flame. There are many defects in these types of fused silica, for example, the air bubbles, inclusions and metallic impurity. The other two types are made by synthetic reaction of SiCl4 with water in oxyhydrogen or plasma flame. Both type Ⅲ and Ⅳ have excellent performance in transmittance and internal quality. However, type Ⅳof fused silica has disadvantage in small aperture and overall high manufacturing cost. Take the transmittance and internal quality into consideration, the type Ⅲ fused silica is the most suitable for large-aperture lens, and can withstand high power laser. The systemic studies of manufacturing process were done to improve the performance of type Ⅲ fused silica in various areas, for instance, the optical homogeneity, the stress birefringence, the absorption coefficient and the damage threshold. There are four steps in manufacturing process of type Ⅲ fused silica, ingot production, reshaping, annealing and cold-working. The critical factors of ingot production, like the flame of burner and the structure of furnace, were deeply studied in this paper to improve the performance of fused silica. On the basis of the above research, the performance and quality of the fused silica measured up to advanced world levels. For instance, the result of optical homogeneity can be controlled to 2-5 ppm, the stress birefringence is better than 4 nm/cm, the absorption coefficient is about 5.971ppm cm-1 (1ω), the damage threshold is greater than 80, 25 and 23 J/cm2 with the wavelength at 1064, 532 and 351nm respectively, the bandwidth used for measuring is 3ns.The fused silica has already been used in the area of high power laser facilities, aerospace industry, primary lens of interferometer based on its excellent performance.

Yeast transcriptional activator INO2 interacts as an Ino2p/Ino4p basic helix-loop-helix heteromeric complex with the inositol/choline-responsive element necessary for expression of phospholipid biosynthetic genes in Saccharomyces cerevisiae.

PubMed Central

Schwank, S; Ebbert, R; Rautenstrauss, K; Schweizer, E; Schüller, H J

1995-01-01

Coordinate transcriptional control of yeast genes involved in phospholipid biosynthesis is mediated by the inositol/choline-responsive element (ICRE) contained in the respective promoter regions. Regulatory genes INO2 and INO4, both encoding basic helix-loop-helix (bHLH) proteins, are necessary for ICRE-dependent gene activation. By the use of size variants and by heterologous expression in E. coli we demonstrate that Ino2p and Ino4p are both necessary and sufficient for the formation of the previously described FAS binding factor 1, Fbf1, interacting with the ICRE. Formation of a heteromeric complex between Ino2p and Ino4p by means of the respective bHLH domains was demonstrated in vivo by the interaction of appropriate two-hybrid constructs and in vitro by Far-Western analyses. Neither Ino2p nor Ino4p binds to the ICRE as a homodimer. When fused to the DNA-binding domain of Gal4p, Ino2p but not Ino4p was able to activate a UASGAL-containing reporter gene even in the absence of the heterologous Fbf1 subunit. By deletion studies, two separate transcriptional activation domains were identified in the N-terminal part of Ino2p. Thus, the bHLH domains of Ino2p and Ino4p constitute the dimerization/DNA-binding module of Fbf1 mediating its interaction with the ICRE, while transcriptional activation is effected exclusively by Ino2p. Images PMID:7862526

The competitor-introduced Ggamma recruitment system, a new approach for screening affinity-enhanced proteins.

PubMed

Fukuda, Nobuo; Ishii, Jun; Tanaka, Tsutomu; Kondo, Akihiko

2010-04-01

We have developed a new approach based on the Ggamma recruitment system to screen affinity-enhanced proteins by expressing a binding competitor. The previously established Ggamma recruitment system is a yeast two-hybrid (Y2H) system that utilizes G-protein signaling, and is based on the fact that membrane localization of the G-protein gamma subunit (Ggamma) is essential for signal transduction in yeast. In the original Y2H system, an engineered Ggamma that lacks membrane localization upon deletion of the lipid modification site (Ggamma(cyto)) is produced, and a candidate protein with an artificial lipidation site and its counterpart fused with Ggamma(cyto) are expressed. As protein-protein interactions bring Ggamma(cyto) towards the plasma membrane, G-protein signaling can be activated, and the interaction is detected by various cellular responses as the readout. In the current study, we expressed a third cytosolic protein that competes with the candidate protein to specifically isolate affinity-enhanced mutants from a mutation library of the candidate protein. Enhancing the affinity of the protein candidate guides the counterpart-Ggamma(cyto) fusion protein towards the plasma membrane and activates signaling. Using mutants of the Z domain derived from Staphylococcus aureus protein A as candidate proteins or competitors, and the Fc portion of human immunoglobulin G (IgG) as the counterpart, we demonstrate that affinity-enhanced proteins can be effectively screened from a library containing a 10 000-fold excess of non-enhanced proteins. This new approach, called the competitor-introduced Ggamma recruitment system, will be useful for efficient discovery of rare valuable candidates hidden among excess ordinary ones.

Mitochondrial anchorage and fusion contribute to mitochondrial inheritance and quality control in the budding yeast Saccharomyces cerevisiae.

PubMed

Higuchi-Sanabria, Ryo; Charalel, Joseph K; Viana, Matheus P; Garcia, Enrique J; Sing, Cierra N; Koenigsberg, Andrea; Swayne, Theresa C; Vevea, Jason D; Boldogh, Istvan R; Rafelski, Susanne M; Pon, Liza A

2016-03-01

Higher-functioning mitochondria that are more reduced and have less ROS are anchored in the yeast bud tip by the Dsl1-family protein Mmr1p. Here we report a role for mitochondrial fusion in bud-tip anchorage of mitochondria. Fluorescence loss in photobleaching (FLIP) and network analysis experiments revealed that mitochondria in large buds are a continuous reticulum that is physically distinct from mitochondria in mother cells. FLIP studies also showed that mitochondria that enter the bud can fuse with mitochondria that are anchored in the bud tip. In addition, loss of fusion and mitochondrial DNA (mtDNA) by deletion of mitochondrial outer or inner membrane fusion proteins (Fzo1p or Mgm1p) leads to decreased accumulation of mitochondria at the bud tip and inheritance of fitter mitochondria by buds compared with cells with no mtDNA. Conversely, increasing the accumulation and anchorage of mitochondria in the bud tip by overexpression of MMR1 results in inheritance of less-fit mitochondria by buds and decreased replicative lifespan and healthspan. Thus quantity and quality of mitochondrial inheritance are ensured by two opposing processes: bud-tip anchorage by mitochondrial fusion and Mmr1p, which favors bulk inheritance; and quality control mechanisms that promote segregation of fitter mitochondria to the bud. © 2016 Higuchi-Sanabria et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Additive manufacturing of glass for optical applications

NASA Astrophysics Data System (ADS)

Luo, Junjie; Gilbert, Luke J.; Bristow, Douglas A.; Landers, Robert G.; Goldstein, Jonathan T.; Urbas, Augustine M.; Kinzel, Edward C.

2016-04-01

Glasses including fused quartz have significant scientific and engineering applications including optics, communications, electronics, and hermetic seals. This paper investigates a filament fed process for Additive Manufacturing (AM) of fused quartz. Additive manufacturing has several potential benefits including increased design freedom, faster prototyping, and lower processing costs for small production volumes. However, current research in AM of glasses is limited and has focused on non-optical applications. Fused quartz is studied here because of its desirability for high-quality optics due to its high transmissivity and thermal stability. Fused quartz also has a higher working temperature than soda lime glass which poses a challenge for AM. In this work, fused quartz filaments are fed into a CO2 laser generated melt pool, smoothly depositing material onto the work piece. Single tracks are printed to explore the effects that different process parameters have on the morphology of printed fused quartz. A spectrometer is used to measure the thermal radiation incandescently emitted from the melt pool. Thin-walls are printed to study the effects of layer-to-layer height. Finally, a 3D fused quartz cube is printed using the newly acquired layer height and polished on each surface. The transmittance and index homogeneity of the polished cube are both measured. These results show that the filament fed process has the potential to print fused quartz with optical transparency and of index of refraction uniformity approaching bulk processed glass.

The canister around the FUSE satellite is removed on the pad at CCAS.

NASA Technical Reports Server (NTRS)

1999-01-01

At Launch Pad 17A, Cape Canaveral Air Station (CCAS), workers begin to remove the canister around the top of the NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe. FUSE is scheduled to be launched from CCAS June 23 aboard a Boeing Delta II rocket.

The canister around the FUSE satellite is removed on the pad at CCAS.

NASA Technical Reports Server (NTRS)

1999-01-01

At Launch Pad 17A, Cape Canaveral Air Station (CCAS), workers oversee the removal of the canister from the top of NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe. FUSE is scheduled to be launched from CCAS June 23 aboard a Boeing Delta II rocket.

Yeasts of the soil – obscure but precious

PubMed Central

2018-01-01

Abstract Pioneering studies performed in the nineteenth century demonstrated that yeasts are present in below‐ground sources. Soils were regarded more as a reservoir for yeasts that reside in habitats above it. Later studies showed that yeast communities in soils are taxonomically diverse and different from those above‐ground. Soil yeasts possess extraordinary adaptations that allow them to survive in a wide range of environmental conditions. A few species are promising sources of yeast oils and have been used in agriculture as potential antagonists of soil‐borne plant pathogens or as plant growth promoters. Yeasts have been studied mainly in managed soils such as vineyards, orchards and agricultural fields, and to a lesser extent under forests and grasslands. Our knowledge of soil yeasts is further biased towards temperate and boreal forests, whereas data from Africa, the Americas and Asia are scarce. Although soil yeast communities are often species‐poor in a single sample, they are more diverse on the biotope level. Soil yeasts display pronounced endemism along with a surprisingly high proportion of currently unidentified species. However, like other soil inhabitants, yeasts are threatened by habitat alterations owing to anthropogenic activities such as agriculture, deforestation and urbanization. In view of the rapid decline of many natural habitats, the study of soil yeasts in undisturbed or low‐managed biotopes is extremely valuable. The purpose of this review is to encourage researchers, both biologists and soil scientists, to include soil yeasts in future studies. PMID:29365211

Diversity and killer activity of yeasts in Malaysian fermented food samples.

PubMed

Lim, S L; Tay, S T

2011-08-01

The biodiversity and the killer activity of yeasts isolated from various types of fermented food in Malaysia were investigated in this study. Of 252 yeasts isolated from 48 fermented food samples in this study, 19 yeast species were identified based on sequence analysis of the ITS1-5.8S-ITS2 partial fragments of the yeasts. A total of 29 (11.5%) of the yeast isolates demonstrated killer activity to at least one Candida species tested in this study; including 22 isolates of Trichosporon asahii, 4 isolates of Pichia anomala, and one isolate each of Pichia norvegensis, Pichia fermentans and Issatchenkia orientalis, respectively. The presence of killer yeasts reflects antagonism that occurs during microbial interaction in the fermented food, whereby certain yeasts produce killer toxins and possibly other toxic substances in competition for limited nutrients and space. The anti-Candida activity demonstrated by killer yeasts in this study should be further explored for development of alternative therapy against candidiasis.

Yeast as a model system for mammalian seven-transmembrane segment receptors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jeansonne, N.E.

1994-05-01

Investigators have used the budding yeast Saccharomyces cerevisiae as a model system in which to study the {beta}-adrenergic receptor, the T-cell receptor pathway, initiation of mammalian DNA replication, initiation of mammalian transcription, secretion, the CDC2 kinase system, cell cycle control, and aging, as well as the function of oncogenes. This list continues to growth with the discovery of an immunoglobulin heavy-chain binding homologue in yeast, an Rb binding protein homologue, and a possible yeast arrestin. Yeast is relatively easy to maintain, to grow, and to genetically manipulate. A single gene can be overexpressed, selectively mutated or deleted from its chromosomalmore » location. In this way, the in vivo function of a gene can be studied. It has become reasonable to consider yeast as a model system for studying the seven transmembrane segments (7-TMS) receptor family. Currently, subtypes of the {beta}-adrenergic receptor are being studied in yeast. The receptor and its G{sub {alpha}}-G-protein, trigger the mating pheromone receptor pathway. This provides a powerful assay for determining receptor function. Studies expressing the muscarinic cholinergic receptor in yeast are underway. The yeast pheromone receptor belongs to this receptor family, sharing sequences and secondary structure homology. An effective strategy has been to identify a yeast pathway or process which is homologous to a mammalian system. The pathway is delineated in yeast, identifying other genetic components. Then yeast genes are used to screen for human homologues of these components. The putative human homologues are then expressed in yeast and in mammalian cells to determine function. When this type of {open_quotes}mixing and matching{close_quotes} works, yeast genetics can be a powerful tool. 115 refs.« less

Yeast for virus research

PubMed Central

Zhao, Richard Yuqi

2017-01-01

Budding yeast (Saccharomyces cerevisiae) and fission yeast (Schizosaccharomyces pombe) are two popular model organisms for virus research. They are natural hosts for viruses as they carry their own indigenous viruses. Both yeasts have been used for studies of plant, animal and human viruses. Many positive sense (+) RNA viruses and some DNA viruses replicate with various levels in yeasts, thus allowing study of those viral activities during viral life cycle. Yeasts are single cell eukaryotic organisms. Hence, many of the fundamental cellular functions such as cell cycle regulation or programed cell death are highly conserved from yeasts to higher eukaryotes. Therefore, they are particularly suited to study the impact of those viral activities on related cellular activities during virus-host interactions. Yeasts present many unique advantages in virus research over high eukaryotes. Yeast cells are easy to maintain in the laboratory with relative short doubling time. They are non-biohazardous, genetically amendable with small genomes that permit genome-wide analysis of virologic and cellular functions. In this review, similarities and differences of these two yeasts are described. Studies of virologic activities such as viral translation, viral replication and genome-wide study of virus-cell interactions in yeasts are highlighted. Impacts of viral proteins on basic cellular functions such as cell cycle regulation and programed cell death are discussed. Potential applications of using yeasts as hosts to carry out functional analysis of small viral genome and to develop high throughput drug screening platform for the discovery of antiviral drugs are presented. PMID:29082230

1 mil gold bond wire study.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huff, Johnathon; McLean, Michael B.; Jenkins, Mark W.

2013-05-01

In microcircuit fabrication, the diameter and length of a bond wire have been shown to both affect the current versus fusing time ratio of a bond wire as well as the gap length of the fused wire. This study investigated the impact of current level on the time-to-open and gap length of 1 mil by 60 mil gold bond wires. During the experiments, constant current was provided for a control set of bond wires for 250ms, 410ms and until the wire fused; non-destructively pull-tested wires for 250ms; and notched wires. The key findings were that as the current increases, themore » gap length increases and 73% of the bond wires will fuse at 1.8A, and 100% of the wires fuse at 1.9A within 60ms. Due to the limited scope of experiments and limited data analyzed, further investigation is encouraged to confirm these observations.« less

Non-destructive evaluation of UV pulse laser-induced damage performance of fused silica optics.

PubMed

Huang, Jin; Wang, Fengrui; Liu, Hongjie; Geng, Feng; Jiang, Xiaodong; Sun, Laixi; Ye, Xin; Li, Qingzhi; Wu, Weidong; Zheng, Wanguo; Sun, Dunlu

2017-11-24

The surface laser damage performance of fused silica optics is related to the distribution of surface defects. In this study, we used chemical etching assisted by ultrasound and magnetorheological finishing to modify defect distribution in a fused silica surface, resulting in fused silica samples with different laser damage performance. Non-destructive test methods such as UV laser-induced fluorescence imaging and photo-thermal deflection were used to characterize the surface defects that contribute to the absorption of UV laser radiation. Our results indicate that the two methods can quantitatively distinguish differences in the distribution of absorptive defects in fused silica samples subjected to different post-processing steps. The percentage of fluorescence defects and the weak absorption coefficient were strongly related to the damage threshold and damage density of fused silica optics, as confirmed by the correlation curves built from statistical analysis of experimental data. The results show that non-destructive evaluation methods such as laser-induced fluorescence and photo-thermal absorption can be effectively applied to estimate the damage performance of fused silica optics at 351 nm pulse laser radiation. This indirect evaluation method is effective for laser damage performance assessment of fused silica optics prior to utilization.

Human adipose tissue-derived stem cells exhibit proliferation potential and spontaneous rhythmic contraction after fusion with neonatal rat cardiomyocytes.

PubMed

Metzele, Roxana; Alt, Christopher; Bai, Xiaowen; Yan, Yasheng; Zhang, Zhi; Pan, Zhizhong; Coleman, Michael; Vykoukal, Jody; Song, Yao-Hua; Alt, Eckhard

2011-03-01

Various types of stem cells have been shown to have beneficial effects on cardiac function. It is still debated whether fusion of injected stem cells with local resident cardiomyocytes is one of the mechanisms. To better understand the role of fusion in stem cell-based myocardial regeneration, the present study was designed to investigate the fate of human adipose tissue-derived stem cells (hASCs) fused with neonatal rat cardiomyocytes in vitro. hASCs labeled with the green fluorescent probe Vybrant DiO were cocultured with neonatal rat cardiomyocytes labeled with the red fluorescent probe Vybrant DiI and then treated with fusion-inducing hemagglutinating virus of Japan (HVJ). Cells that incorporated both red and green fluorescent signals were considered to be hASCs that had fused with rat cardiomyocytes. Fusion efficiency was 19.86 ± 4.84% at 5 d after treatment with HVJ. Most fused cells displayed cardiomyocyte-like morphology and exhibited spontaneous rhythmic contraction. Both immunofluorescence staining and lentiviral vector labeling showed that fused cells contained separate rat cardiomyocyte and hASC nuclei. Immunofluorescence staining assays demonstrated that human nuclei in fused cells still expressed the proliferation marker Ki67. In addition, hASCs fused with rat cardiomyocytes were positive for troponin I. Whole-cell voltage-clamp analysis demonstrated action potentials in beating fused cells. RT-PCR analysis using rat- or human-specific myosin heavy chain primers revealed that the myosin heavy-chain expression in fused cells was derived from rat cardiomyocytes. Real-time PCR identified expression of human troponin T in fused cells and the presence of rat cardiomyocytes induced a cardiomyogenic protein expression of troponin T in human ASCs. This study illustrates that hASCs exhibit both stem cell (proliferation) and cardiomyocyte properties (action potential and spontaneous rhythmic beating) after fusion with rat cardiomyocytes, supporting the theory that fusion, even if artificially induced in our study, could indeed be a mechanism for cardiomyocyte renewal in the heart.

The Influence of Heating Mains on Yeast Communities in Urban Soils

NASA Astrophysics Data System (ADS)

Tepeeva, A. N.; Glushakova, A. M.; Kachalkin, A. V.

2018-04-01

The number and species diversity of yeasts in urban soils (urbanozems) affected by heating mains and in epiphytic yeast complexes of grasses growing above them were studied. The number of yeasts in the soil reached 103-104 CFU/g; on the plants, 107 CFU/g. Significant (by an order of magnitude) increase in the total number of soil yeasts in the zone of heating mains in comparison with the surrounding soil was found in winter period. Overall, 25 species of yeasts were isolated in our study. Yeast community of studied urbanozems was dominated by the Candida sake, an eurybiont of the temperate zone and other natural ecotopes with relatively low temperatures, but its share was minimal in the zone of heating mains. In general, the structure of soil and epiphytic yeast complexes in the zones of heating mains differed from that in the surrounding area by higher species diversity and a lower share of pigmented species among the epiphytic yeasts. The study demonstrated that the number and species structure of soil yeast communities in urban soils change significantly under the influence of the temperature factor and acquire a mosaic distribution pattern.

Dibenzo[a,f]perylene bisimide: effects of introducing two fused rings.

PubMed

Chaolumen; Enno, Hiroki; Murata, Michihisa; Wakamiya, Atsushi; Murata, Yasujiro

2014-11-01

Perylene bisimides (PBIs) are fascinating dyes with various potential applications. To study the effects of introducing a dibenzo-fused structure to the perylene moiety, π-extended PBI derivatives with a dibenzo-fused structure at both of the a and f bonds were synthesized. The twisted structure was characterized by X-ray crystal structure analysis. In the cyclic voltammograms, the dibenzo[a,f]-fused PBI showed a reversible oxidation wave at much less positive potential, relative to a dibenzo[a,o]-fused PBI derivative. These data indicated that two ring fusions at both sides of a naphthalene moiety, which construct a tetracene core, effectively raise the HOMO level compared to fusion of one ring at each naphthalene moiety (two anthracene cores). The dibenzo[a,f]-fused PBI derivative showed an absorption band at 735 nm with a shoulder band reaching 900 nm. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Comparing monolithic and fused core HPLC columns for fast chromatographic analysis of fat-soluble vitamins.

PubMed

Kurdi, Said El; Muaileq, Dina Abu; Alhazmi, Hassan A; Bratty, Mohammed Al; Deeb, Sami El

2017-06-27

HPLC stationary phases of monolithic and fused core type can be used to achieve fast chromatographic separation as an alternative to UPLC. In this study, monolithic and fused core stationary phases are compared for fast separation of four fat-soluble vitamins. Three new methods on the first and second generation monolithic silica RP-18e columns and a fused core pentafluoro-phenyl propyl column were developed. Application of three fused core columns offered comparable separations of retinyl palmitate, DL-α-tocopheryl acetate, cholecalciferol and menadione in terms of elution speed and separation efficiency. Separation was achieved in approx. 5 min with good resolution (Rs > 5) and precision (RSD ≤ 0.6 %). Monolithic columns showed, however, a higher number of theoretical plates, better precision and lower column backpressure than the fused core column. The three developed methods were successfully applied to separate and quantitate fat-soluble vitamins in commercial products.

KSC-99pp0701

NASA Image and Video Library

1999-06-17

At Launch Pad 17A, Cape Canaveral Air Station (CCAS), workers remove another section of the canister surrounding NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe. FUSE is scheduled to be launched from CCAS June 23 aboard a Boeing Delta II rocket

Fluorescence lifetime dynamics of eGFP in protein aggregates with expanded polyQ

NASA Astrophysics Data System (ADS)

Ghukasyan, Vladimir; Hsu, Chih-Chun; Liu, Chia-Rung; Kao, Fu-Jen; Cheng, Tzu-Hao

2009-02-01

Expanding a polyglutamine (polyQ) stretch at the N-terminus of huntingtin protein is the main cause of the neurodegenerative disorder Huntington's disease (HD). Expansion of polyQ above 39 residues has an inherent propensity to form amyloid-like fibrils and aggregation of the mutant protein is found to be a critical component for abnormal pathology of HD. Using yeast Saccharomyces cerevisiae as a model system, we have observed a decrease in fluorescence lifetime of the enhanced green fluorescence protein (eGFP) fused to 97 successive glutamine residues (97Q). Compared to the sample expressing evenly distributed eGFP, the 97Q-eGFP fusion proteins show the formation of grain-like particles and the reduction of eGFP lifetime by ~250 ps as measured by time-correlated single-photon counting technique (TCSPC). More importantly, this phenomenon does not appear in Hsp104-deficient cells. The gene product of HSP104 is required for the formation of polyQ aggregates in yeast cells; therefore, the cellular 97Q-eGFP become soluble and evenly distributive in the absence of Hsp104. Under this condition, the lifetime value of 97Q-eGFP is close to the one exhibited by eGFP alone. The independence of the effect of the environmental parameters, such as pH and refraction index is demonstrated. These data indicate that the fluorescence lifetime dynamics of eGFP is linked to the process of polyQ protein aggregation per se.

Mitochondrial genome maintenance: roles for nuclear nonhomologous end-joining proteins in Saccharomyces cerevisiae.

PubMed

Kalifa, Lidza; Quintana, Daniel F; Schiraldi, Laura K; Phadnis, Naina; Coles, Garry L; Sia, Rey A; Sia, Elaine A

2012-03-01

Mitochondrial DNA (mtDNA) deletions are associated with sporadic and inherited diseases and age-associated neurodegenerative disorders. Approximately 85% of mtDNA deletions identified in humans are flanked by short directly repeated sequences; however, mechanisms by which these deletions arise are unknown. A limitation in deciphering these mechanisms is the essential nature of the mitochondrial genome in most living cells. One exception is budding yeast, which are facultative anaerobes and one of the few organisms for which directed mtDNA manipulation is possible. Using this model system, we have developed a system to simultaneously monitor spontaneous direct-repeat-mediated deletions (DRMDs) in the nuclear and mitochondrial genomes. In addition, the mitochondrial DRMD reporter contains a unique KpnI restriction endonuclease recognition site that is not present in otherwise wild-type (WT) mtDNA. We have expressed KpnI fused to a mitochondrial localization signal to induce a specific mitochondrial double-strand break (mtDSB). Here we report that loss of the MRX (Mre11p, Rad50p, Xrs2p) and Ku70/80 (Ku70p, Ku80p) complexes significantly impacts the rate of spontaneous deletion events in mtDNA, and these proteins contribute to the repair of induced mtDSBs. Furthermore, our data support homologous recombination (HR) as the predominant pathway by which mtDNA deletions arise in yeast, and suggest that the MRX and Ku70/80 complexes are partially redundant in mitochondria.

Mitochondrial Genome Maintenance: Roles for Nuclear Nonhomologous End-Joining Proteins in Saccharomyces cerevisiae

PubMed Central

Kalifa, Lidza; Quintana, Daniel F.; Schiraldi, Laura K.; Phadnis, Naina; Coles, Garry L.; Sia, Rey A.; Sia, Elaine A.

2012-01-01

Mitochondrial DNA (mtDNA) deletions are associated with sporadic and inherited diseases and age-associated neurodegenerative disorders. Approximately 85% of mtDNA deletions identified in humans are flanked by short directly repeated sequences; however, mechanisms by which these deletions arise are unknown. A limitation in deciphering these mechanisms is the essential nature of the mitochondrial genome in most living cells. One exception is budding yeast, which are facultative anaerobes and one of the few organisms for which directed mtDNA manipulation is possible. Using this model system, we have developed a system to simultaneously monitor spontaneous direct-repeat–mediated deletions (DRMDs) in the nuclear and mitochondrial genomes. In addition, the mitochondrial DRMD reporter contains a unique KpnI restriction endonuclease recognition site that is not present in otherwise wild-type (WT) mtDNA. We have expressed KpnI fused to a mitochondrial localization signal to induce a specific mitochondrial double-strand break (mtDSB). Here we report that loss of the MRX (Mre11p, Rad50p, Xrs2p) and Ku70/80 (Ku70p, Ku80p) complexes significantly impacts the rate of spontaneous deletion events in mtDNA, and these proteins contribute to the repair of induced mtDSBs. Furthermore, our data support homologous recombination (HR) as the predominant pathway by which mtDNA deletions arise in yeast, and suggest that the MRX and Ku70/80 complexes are partially redundant in mitochondria. PMID:22214610

The cel3 gene of Agaricus bisporus codes for a modular cellulase and is transcriptionally regulated by the carbon source.

PubMed Central

Chow, C M; Yagüe, E; Raguz, S; Wood, D A; Thurston, C F

1994-01-01

A 52-kDa protein, CEL3, has been separated from the culture filtrate of Agaricus bisporus during growth on cellulose. A PCR-derived probe was made, with a degenerate oligodeoxynucleotide derived from the amino acid sequence of a CEL3 CNBr cleavage product and was used to select cel3 cDNA clones from an A. bisporus cDNA library. Two allelic cDNAs were isolated. They showed 98.8% identity of their nucleotide sequences. The deduced amino acid sequence and domain architecture of CEL3 showed a high degree of similarity to those of cellobiohydrolase II of Trichoderma reesei. Functional expression of cel3 cDNA in Saccharomyces cerevisiae was achieved by placing it under the control of a constitutive promoter and fusing it to the yeast invertase signal sequence. Recombinant CEL3 secreted by yeast showed enzymatic activity towards crystalline cellulose. At long reaction times, CEL3 was also able to degrade carboxymethyl cellulose. Northern (RNA) analysis showed that cel3 gene expression was induced by cellulose and repressed by glucose, fructose, 2-deoxyglucose, and lactose. Glycerol, mannitol, sorbitol, and maltose were neutral carbon sources. Nuclear run-on analysis showed that the rate of synthesis of cel3 mRNA in cellulose-grown cultures was 13 times higher than that in glucose-grown cultures. A low basal rate of cel3 mRNA synthesis was observed in the nuclei isolated from glucose-grown mycelia. Images PMID:8085821

Cell permeability and nuclear DNA staining by propidium iodide in basidiomycetous yeasts.

PubMed

Zhang, Ning; Fan, Yuxuan; Li, Chen; Wang, Qiming; Leksawasdi, Noppol; Li, Fuli; Wang, Shi'an

2018-05-01

Non-model yeasts within basidiomycetes have considerable importance in agriculture, industry, and environment, but they are not as well studied as ascomycetous yeasts. Serving as a basic technique, nuclear DNA staining is widely used in physiology, ecology, cell biology, and genetics. However, it is unclear whether the classical nuclear DNA staining method for ascomycetous yeasts is applicable to basidiomycetous yeasts. In this study, 5 yeasts ineffectively stained by the classical propidium iodide (PI) staining method were identified from 23 representative basidiomycetous yeasts. Pretreatment of cells using dimethyl sulfoxide (DMSO) or snailase markedly improved cell penetration to PI and thus enabled DNA content determination by flow cytometry on the recalcitrant yeasts. The pretreatments are efficient, simple, and fast, avoiding tedious mutagenesis or genetic engineering used in previous reports. The heterogeneity of cell penetration to PI among basidiomycetous yeasts was attributed to the discrepancy in cell wall polysaccharides instead of capsule or plasma membrane. This study also indicated that care must be taken in attributing PI-negative staining as viable cells when studying non-model microorganisms.

[Study on mechanism of inactivated cider yeast adsorbing patulin by Fourier transform infrared spectroscopy].

PubMed

Guo, Cai-Xia; Yue, Tian-Li; Yuan, Ya-Hong; Wang, Zhou-Li; Wang, Ling; Cai, Rui

2013-03-01

The mechanism of patulin adsorption by inactivated cider yeast was studied by chemical modification and FTIR The results of patulin removal by various modified yeast biomass showed that the ability of patulin biosorption by acetone-treated yeast and NaOH-treated yeast increased siginificantly, while the methylation of amino group and esterification of carboxylate functionalities of yeast cell surface caused a decrease in patulin binding, which indicated that amino group and carboxyl group presented in the cell walls of yeast might be involved in the binding of patulin to the yeast. The FTIR analysis indicated that the main functional groups were amino group, carboxyl group and hydroxy group which are associated with protein and polysaccharides.

Yeast Modulation of Human Dendritic Cell Cytokine Secretion: An In Vitro Study

PubMed Central

Smith, Ida M.; Christensen, Jeffrey E.; Arneborg, Nils; Jespersen, Lene

2014-01-01

Probiotics are live microorganisms which when administered in adequate amounts confer a health benefit on the host. The concept of individual microorganisms influencing the makeup of T cell subsets via interactions with intestinal dendritic cells (DCs) appears to constitute the foundation for immunoregulatory effects of probiotics, and several studies have reported probiotic strains resulting in reduction of intestinal inflammation through modulation of DC function. Consequent to a focus on Saccharomyces boulardii as the fundamental probiotic yeast, very little is known about hundreds of non-Saccharomyces yeasts in terms of their interaction with the human gastrointestinal immune system. The aim of the present study was to evaluate 170 yeast strains representing 75 diverse species for modulation of inflammatory cytokine secretion by human DCs in vitro, as compared to cytokine responses induced by a S. boulardii reference strain with probiotic properties documented in clinical trials. Furthermore, we investigated whether cytokine inducing interactions between yeasts and human DCs are dependent upon yeast viability or rather a product of membrane interactions regardless of yeast metabolic function. We demonstrate high diversity in yeast induced cytokine profiles and employ multivariate data analysis to reveal distinct clustering of yeasts inducing similar cytokine profiles in DCs, highlighting clear species distinction within specific yeast genera. The observed differences in induced DC cytokine profiles add to the currently very limited knowledge of the cross-talk between yeasts and human immune cells and provide a foundation for selecting yeast strains for further characterization and development toward potentially novel yeast probiotics. Additionally, we present data to support a hypothesis that the interaction between yeasts and human DCs does not solely depend on yeast viability, a concept which may suggest a need for further classifications beyond the current definition of a probiotic. PMID:24816850

Additive manufacturing of transparent fused quartz

NASA Astrophysics Data System (ADS)

Luo, Junjie; Hostetler, John M.; Gilbert, Luke; Goldstein, Jonathan T.; Urbas, Augustine M.; Bristow, Douglas A.; Landers, Robert G.; Kinzel, Edward C.

2018-04-01

This paper investigates a filament-fed process for additive manufacturing (AM) of fused quartz. Glasses such as fused quartz have significant scientific and engineering applications, which include optics, communications, electronics, and hermetic seals. AM has several attractive benefits such as increased design freedom, faster prototyping, and lower processing costs for small production volumes. However, current research into glass AM has focused primarily on nonoptical applications. Fused quartz is studied here because of its desirability for use in high-quality optics due to its high transmissivity and thermal stability. Fused quartz filaments are fed into a CO2 laser-generated molten region, smoothly depositing material onto the workpiece. Spectroscopy and pyrometry are used to measure the thermal radiation incandescently emitted from the molten region. The effects of the laser power and scan speed are determined by measuring the morphology of single tracks. Thin walls are printed to study the effects of layer-to-layer height. This information is used to deposit solid pieces including a cylindrical-convex shape capable of focusing visible light. The transmittance and index homogeneity of the printed fused quartz are measured. These results show that the filament-fed process has the potential to print transmissive optics.

[Study on content of nickel in saliva released from the nickel-chromium and the nickel-chromium-titanium porcelain alloy].

PubMed

Wang, Wen-Jie; Zhang, Tai-Qiang; Wei, Hong

2010-02-01

The purpose of the study was to investigate the content of nickel (Ni) ion in patients' saliva after wearing the porcelain-fused-to nickel-chromium (Ni-Cr) crown or the porcelain-fused-to nickel-chromium-titanium(Ni-Cr-Ti) crown. 50 patients who had one molar or premolar needed repairing were selected and divided into two groups randomly. Patients in one group were fabricated with porcelain-fused-to Ni-Cr crown and the patients in the other group were fabricated with porcelain-fused-to Ni-Cr-Ti crown. Collect the patients' saliva before wearing, 1 week, 3 months, and 6 months after wearing. The content of Ni ion in saliva was detected by inductively coupled plasma mass spectrometry (ICP-MS). The content of Ni ion in both groups increased at the first week, and go back after 6 months. There were no significant differences before wearing, 1 week, 3 months, and 6 months after wearing. There were no significant differences between the two groups. Wearing the porcelain-fused-to Ni-Cr crown or the porcelain-fused-to Ni-Cr-Ti crown has no significant influence on the content of Ni ion in saliva.

Effect of wine yeast monoculture practice on the biodiversity of non-Saccharomyces yeasts.

PubMed

Ganga, M A; Martínez, C

2004-01-01

The objective of this work was to study the effect of the use of Saccharomyces cerevisiae monocultures over the biodiversity of non-Saccharomyces yeasts in wine-producing areas in Chile. Microvinifications were carried out with grape musts of two areas. In one of them, the fermentation is carried out mainly in a spontaneous manner, whereas in the other the musts are inoculated with commercial yeasts. The isolated yeasts were identified by the internal transcribed (ITS)/restriction fragment length polymorphism technique. In the industrial production area less variability of yeast genera was observed as compared with the traditional area, an observation that is greatest at the end of the fermentation. Furthermore, a study of the production of extracellular enzymes was done. The majority of the yeasts showed at least one of the activities assayed with the exception of beta-glycosidase. The results suggest that in the industrialized area the diversity of yeasts is less in the traditional area. Likewise, the potentiality of the non-Saccharomyces yeasts as enzyme producers with industrial interest has been confirmed. This study shows the negative effect of the use of monocultures over the biodiversity of yeasts in wine-producing regions.

Glycosylatable GFP as a compartment-specific membrane topology reporter

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Hunsang; Min, Jisoo; Heijne, Gunnar von

2012-11-02

Highlights: Black-Right-Pointing-Pointer An N-linked glycosylation site is introduced near the GFP fluorophore. Black-Right-Pointing-Pointer gGFP is not glycosylated and is fully fluorescent in the cytosol. Black-Right-Pointing-Pointer gGFP is glycosylated and non-fluorescent in the lumen of the ER. Black-Right-Pointing-Pointer gGFP is fused to membrane proteins of known topology. Black-Right-Pointing-Pointer Its applicability as a membrane topology reporter is demonstrated. -- Abstract: Determination of the membrane topology is an essential step in structural and functional studies of integral membrane proteins, yet the choices of membrane topology reporters are limited and the experimental analysis can be laborious, especially in eukaryotic cells. Here, we present amore » robust membrane topology reporter, glycosylatable green fluorescent protein (gGFP). gGFP is fully fluorescent in the yeast cytosol but becomes glycosylated and does not fluoresce in the lumen of the endoplasmic reticulum (ER). Thus, by assaying fluorescence and the glycosylation status of C-terminal fusions of gGFP to target membrane proteins in whole-cell lysates, the localization of the gGFP moiety (and hence the fusion joint) relative to the ER membrane can be unambiguously determined.« less

Identifying Protein-protein Interaction in Drosophila Adult Heads by Tandem Affinity Purification (TAP)

PubMed Central

Tian, Xiaolin; Zhu, Mingwei; Li, Long; Wu, Chunlai

2013-01-01

Genetic screens conducted using Drosophila melanogaster (fruit fly) have made numerous milestone discoveries in the advance of biological sciences. However, the use of biochemical screens aimed at extending the knowledge gained from genetic analysis was explored only recently. Here we describe a method to purify the protein complex that associates with any protein of interest from adult fly heads. This method takes advantage of the Drosophila GAL4/UAS system to express a bait protein fused with a Tandem Affinity Purification (TAP) tag in fly neurons in vivo, and then implements two rounds of purification using a TAP procedure similar to the one originally established in yeast1 to purify the interacting protein complex. At the end of this procedure, a mixture of multiple protein complexes is obtained whose molecular identities can be determined by mass spectrometry. Validation of the candidate proteins will benefit from the resource and ease of performing loss-of-function studies in flies. Similar approaches can be applied to other fly tissues. We believe that the combination of genetic manipulations and this proteomic approach in the fly model system holds tremendous potential for tackling fundamental problems in the field of neurobiology and beyond. PMID:24335807

Data fusion in data scarce areas using a back-propagation artificial neural network model: a case study of the South China Sea

NASA Astrophysics Data System (ADS)

Wang, Zheng; Mao, Zhihua; Xia, Junshi; Du, Peijun; Shi, Liangliang; Huang, Haiqing; Wang, Tianyu; Gong, Fang; Zhu, Qiankun

2018-06-01

The cloud cover for the South China Sea and its coastal area is relatively large throughout the year, which limits the potential application of optical remote sensing. A HJ-charge-coupled device (HJ-CCD) has the advantages of wide field, high temporal resolution, and short repeat cycle. However, this instrument suffers from its use of only four relatively low-quality bands which can't adequately resolve the features of long wavelengths. The Landsat Enhanced Thematic Mapper-plus (ETM+) provides high-quality data, however, the Scan Line Corrector (SLC) stopped working and caused striping of remote sensed images, which dramatically reduced the coverage of the ETM+ data. In order to combine the advantages of the HJ-CCD and Landsat ETM+ data, we adopted a back-propagation artificial neural network (BP-ANN) to fuse these two data types for this study. The results showed that the fused output data not only have the advantage of data intactness for the HJ-CCD, but also have the advantages of the multi-spectral and high radiometric resolution of the ETM+ data. Moreover, the fused data were analyzed qualitatively, quantitatively and from a practical application point of view. Experimental studies indicated that the fused data have a full spatial distribution, multi-spectral bands, high radiometric resolution, a small difference between the observed and fused output data, and a high correlation between the observed and fused data. The excellent performance in its practical application is a further demonstration that the fused data are of high quality.

Morphological evaluation of maxillary second molars with fused roots: a micro-CT study.

PubMed

Ordinola-Zapata, R; Martins, J N R; Bramante, C M; Villas-Boas, M H; Duarte, M H; Versiani, M A

2017-12-01

To evaluate the internal and external morphologies of fused-rooted maxillary second molars by means of micro-computed tomography (micro-CT) analysis. A total of 100 fused-rooted maxillary second molars from a Brazilian subpopulation were divided into six groups according to the root morphology. The samples were scanned at a resolution of 19.6 μm and evaluated with regard to the external morphology of the roots, the root canal configuration, the percentage frequency of C-shaped canals and isthmuses, as well as the morphology of the root canal system at 1, 2 and 3 mm from the anatomical apex of the fused roots. The most prevalent root canal fusions were type 1, mesiobuccal root fused with distobuccal root (32%), followed by type 3, DB root fused with P root (27%), and type 4, MB root fused with DB root, and P root fused with MB or DB roots (21%). The prevalence of C-shaped root canal systems were 22%. Depending on the type of root fusion, the percentage frequency of isthmuses in the apical level varied from 9.3% to 42.8%, whilst the presence of apical deltas ranged from 18.5% to 57.1% of teeth. The root canal system of maxillary second molars with fused roots may have a high incidence of merging canals, isthmuses, apical deltas and C-shaped configurations. © 2017 International Endodontic Journal. Published by John Wiley & Sons Ltd.

KSC-99pp0700

NASA Image and Video Library

1999-06-17

At Launch Pad 17A, Cape Canaveral Air Station (CCAS), workers begin removing the lower sections of the canister surrounding NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe. FUSE is scheduled to be launched from CCAS June 23 aboard a Boeing Delta II rocket

KSC-99pp0699

NASA Image and Video Library

1999-06-17

At Launch Pad 17A, Cape Canaveral Air Station (CCAS), workers oversee the removal of the canister from the top of NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe. FUSE is scheduled to be launched from CCAS June 23 aboard a Boeing Delta II rocket

KSC-99pp0698

NASA Image and Video Library

1999-06-17

At Launch Pad 17A, Cape Canaveral Air Station (CCAS), workers begin to remove the canister around the top of the NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe. FUSE is scheduled to be launched from CCAS June 23 aboard a Boeing Delta II rocket

KSC-99pp0703

NASA Image and Video Library

1999-06-17

At Launch Pad 17A, Cape Canaveral Air Station (CCAS), workers look over NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite after sections of the canister have been removed. FUSE is scheduled to be launched from CCAS June 23 aboard a Boeing Delta II rocket. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe

[Color stability of porcelain-fused-to-titanium restorations after repeated firings].

PubMed

Wu, Xiao-min; Zhou, Zheng

2004-08-01

To study the effect of repeated firings on color of porcelain-fused-to-titanium restorations. Twenty standard porcelain-fused-to-titanium specimens were made with Dentsply DETREY TiBond C3 and Dentaurum rematitan Til. Then they were fired 1, 2, 4, 7, and 10 times. Color measurement was done after each firing. Most of the color parameters had no significant difference after firing except for a* and Hab. All the color difference among groups were smaller than 1.0 (deltaE < 1.0). Repeated firings will not affect the color stability of porcelain-fused-to-titanium restorations.

Two soybean bHLH factors regulate response to iron deficiency.

PubMed

Li, Lin; Gao, Wenwen; Peng, Qi; Zhou, Bin; Kong, Qihui; Ying, Yinghui; Shou, Huixia

2018-03-25

Iron is an indispensable micronutrient for plant growth and development. Limited bioavailability of Fe in the soil leads to iron deficiency chlorosis in plants and yield loss. In this study, two soybean basic helix-loop-helix transcription factors, GmbHLH57 and GmbHLH300, were identified in response to Fe-deficiency. Both transcription factors are expressed in roots and nodules, and are induced by Fe deficiency; these patterns were confirmed in transgenic hairy roots expressing constructs of the endogenous promoters fused to a GUS reporter gene. Bimolecular fluorescence complementation, yeast two-hybrid and coimmunoprecipitation (co-IP) assays indicated a physical interaction between GmbHLH57 and GmbHLH300. Studies on transgenic soybeans overexpressing GmbHLH57 and GmbHLH300 revealed that overexpression of each transcription factor, alone, results in no change of the responses to Fe deficiency, whereas overexpression of both transcription factors upregulated the downstream Fe uptake genes and increased the Fe content in these transgenic plants. Compared to wild type, these double overexpression transgenic plants were more tolerant to Fe deficiency. Taken together, our findings establish that GmbHLH57 and GmbHLH300 are important transcription factors involved in Fe homeostasis in soybean. © 2018 Institute of Botany, Chinese Academy of Sciences.

Expression, purification, and antibacterial activity of bovine lactoferrampin-lactoferricin in Pichia pastoris.

PubMed

Tang, Xiang-Shan; Tang, Zhi-Ru; Wang, Sheng-Ping; Feng, Ze-Meng; Zhou, Dong; Li, Tie-Jun; Yin, Yu-Long

2012-02-01

Bovine lactoferrampin (LFA) and bovine lactoferricin (LFC) are two antimicrobial peptides located in the N(1) domain of bovine lactoferrin. The bactericidal activity of the fused peptide LFA-LFC is stronger than that of either LFA or LFC. The high cost of peptide production from either native digestion or chemical synthesis limits the clinical application of antimicrobial peptides. The expression of recombinant peptides in yeast may be an effective alternative. In the current study, the expression, purification, and antibacterial activity of LFA-LFC using the Pichia pastoris expression system are reported. The linearized expression vector pPICZaA-LFA-LFC was transformed into P. pastoris KM71 by electroporation, and positive colonies harboring the target genes were screened out and used for fermentation. The recombinant LFA-LFC peptide was purified via two-step column chromatography and identified by tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The results indicate that P. pastoris is a suitable system for secreting LFA-LFC. The fermentation supernate and the purified LFA-LFC show high antimicrobial activities. The current study is the first to report on the expression and purification of LFA-LFC in P. pastoris and may have potential practical applications in microbial peptide production.

Chemical signaling and insect attraction is a conserved trait in yeasts.

PubMed

Becher, Paul G; Hagman, Arne; Verschut, Vasiliki; Chakraborty, Amrita; Rozpędowska, Elżbieta; Lebreton, Sébastien; Bengtsson, Marie; Flick, Gerhard; Witzgall, Peter; Piškur, Jure

2018-03-01

Yeast volatiles attract insects, which apparently is of mutual benefit, for both yeasts and insects. However, it is unknown whether biosynthesis of metabolites that attract insects is a basic and general trait, or if it is specific for yeasts that live in close association with insects. Our goal was to study chemical insect attractants produced by yeasts that span more than 250 million years of evolutionary history and vastly differ in their metabolism and lifestyle. We bioassayed attraction of the vinegar fly Drosophila melanogaster to odors of phylogenetically and ecologically distinct yeasts grown under controlled conditions. Baker's yeast Saccharomyces cerevisiae , the insect-associated species Candida californica , Pichia kluyveri and Metschnikowia andauensis , wine yeast Dekkera bruxellensis , milk yeast Kluyveromyces lactis , the vertebrate pathogens Candida albicans and Candida glabrata , and oleophilic Yarrowia lipolytica were screened for fly attraction in a wind tunnel. Yeast headspace was chemically analyzed, and co-occurrence of insect attractants in yeasts and flowering plants was investigated through a database search. In yeasts with known genomes, we investigated the occurrence of genes involved in the synthesis of key aroma compounds. Flies were attracted to all nine yeasts studied. The behavioral response to baker's yeast was independent of its growth stage. In addition to Drosophila , we tested the basal hexapod Folsomia candida (Collembola) in a Y-tube assay to the most ancient yeast, Y. lipolytica, which proved that early yeast signals also function on clades older than neopteran insects. Behavioral and chemical data and a search for selected genes of volatile metabolites underline that biosynthesis of chemical signals is found throughout the yeast clade and has been conserved during the evolution of yeast lifestyles. Literature and database reviews corroborate that yeast signals mediate mutualistic interactions between insects and yeasts. Moreover, volatiles emitted by yeasts are commonly found also in flowers and attract many insect species. The collective evidence suggests that the release of volatile signals by yeasts is a widespread and phylogenetically ancient trait, and that insect-yeast communication evolved prior to the emergence of flowering plants. Co-occurrence of the same attractant signals in yeast and flowers suggests that yeast-insect communication may have contributed to the evolution of insect-mediated pollination in flowers.

Practical comparison of 2.7 microm fused-core silica particles and porous sub-2 microm particles for fast separations in pharmaceutical process development.

PubMed

Abrahim, Ahmed; Al-Sayah, Mohammad; Skrdla, Peter; Bereznitski, Yuri; Chen, Yadan; Wu, Naijun

2010-01-05

Fused-core silica stationary phases represent a key technological advancement in the arena of fast HPLC separations. These phases are made by fusing a 0.5 microm porous silica layer onto 1.7 microm nonporous silica cores. The reduced intra-particle flow path of the fused particles provides superior mass transfer kinetics and better performance at high mobile phase velocities, while the fused-core particles provide lower pressure than sub-2 microm particles. In this work, chromatographic performance of the fused-core particles (Ascentis Express) was investigated and compared to that of sub-2 microm porous particles (1.8 microm Zorbax Eclipse Plus C18 and 1.7 microm Acquity BEH C18). Specifically, retention, selectivity, and loading capacity were systematically compared for these two types of columns. Other chromatographic parameters such as efficiency and pressure drop were also studied. Although the fused-core column was found to provide better analyte shape selectivity, both columns had similar hydrophobic, hydrogen bonding, total ion-exchange, and acidic ion-exchange selectivities. As expected, the retention factors and sample loading capacity on the fused-core particle column were slightly lower than those for the sub-2 microm particle column. However, the most dramatic observation was that similar efficiency separations to the sub-2 microm particles could be achieved using the fused-core particles, without the expense of high column back pressure. The low pressure of the fused-core column allows fast separations to be performed routinely on a conventional LC system without significant loss in efficiency or resolution. Applications to the HPLC impurity profiling of drug substance candidates were performed using both types of columns to validate this last point.

Redundancy Technology With A Focused Ion Beam

NASA Astrophysics Data System (ADS)

Komano, Haruki; Hashimoto, Kazuhiko; Takigawa, Tadahiro

1989-08-01

Fuse cutting with a focused ion beam to activate redundancy circuits is proposed. In order to verify its potential usefulness, experiments have been performed. Fuse-cutting time was evaluated using aluminum fuses with a thin passivation layer, which are difficult to cut by conventional laser-beam technology due to the material's high reflectivity. The fuse width and thickness were 2 and 0.8 μm, respectively. The fuse was cut in 5 seconds with a 30 keV focused ion beam of 0.3 A/cm2 current density. Since the fuses used in DRAMs will be smaller, their cutting time will become shorter by scanning an ion beam on narrower areas. Moreover, it can be shortened by increasing current density. Fuses for redundancy technology in 256 k CMOS SRAMs were cut with a focused ion beam. The operation of the memories was checked with a memory tester. It was confirmed that memories which had failure cells operated normally after focused-ion-beam fuse-cutting. Focused ion beam irradiation effects upon a device have been studied. When a 30 keV gallium focused ion beam was irradiated near the gate of MOSFETs, a threshold voltage shift was not observed at an ion dose of 0.3 C/cm2 which corresponded to the ion dose in cutting a fuse. However, when irradiated on the gate, a threshold voltage shift was observed at ion doses of more than 8 x 10-4 C/cm2. The voltage shift was caused by the charge of ions within the passivation layer. It is necessary at least not to irradiate a focused ion beam on a device in cutting fuses. It is concluded that the focused-ion-beam method will be advantageous for future redundancy technology application.

Nectar sugars and bird visitation define a floral niche for basidiomycetous yeast on the Canary Islands.

PubMed

Mittelbach, Moritz; Yurkov, Andrey M; Nocentini, Daniele; Nepi, Massimo; Weigend, Maximilian; Begerow, Dominik

2015-02-01

Studies on the diversity of yeasts in floral nectar were first carried out in the late 19th century. A narrow group of fermenting, osmophilous ascomycetes were regarded as exclusive specialists able to populate this unique and species poor environment. More recently, it became apparent that microorganisms might play an important role in the process of plant pollination. Despite the importance of these nectar dwelling yeasts, knowledge of the factors that drive their diversity and species composition is scarce. In this study, we linked the frequencies of yeast species in floral nectars from various host plants on the Canary Islands to nectar traits and flower visitors. We estimated the structuring impact of pollination syndromes (nectar volume, sugar concentration and sugar composition) on yeast diversity.The observed total yeast diversity was consistent with former studies, however, the present survey yielded additional basidiomycetous yeasts in unexpectedly high numbers. Our results show these basidiomycetes are significantly associated with ornithophilous flowers. Specialized ascomycetes inhabit sucrose-dominant nectars, but are surprisingly rare in nectar dominated by monosaccharides. There are two conclusions from this study: (i) a shift of floral visitors towards ornithophily alters the likelihood of yeast inoculation in flowers, and (ii) low concentrated hexose-dominant nectar promotes colonization of flowers by basidiomycetes. In the studied floral system, basidiomycete yeasts are acknowledged as regular members of nectar. This challenges the current understanding that nectar is an ecological niche solely occupied by ascomycetous yeasts.

Adenoviral-Mediated Imaging of Gene Transfer Using a Somatostatin Receptor-Cytosine Deaminase Fusion Protein

PubMed Central

Lears, Kimberly A.; Parry, Jesse J.; Andrews, Rebecca; Nguyen, Kim; Wadas, Thaddeus J.; Rogers, Buck E.

2015-01-01

Suicide gene therapy is a process by which cells are administered a gene that encodes a protein capable of converting a nontoxic prodrug into an active toxin. Cytosine deaminase (CD) has been widely investigated as a means of suicide gene therapy due to the enzyme’s ability to convert the prodrug 5-fluorocytosine (5-FC) into the toxic compound 5-fluorouracil (5-FU). However, the extent of gene transfer is a limiting factor in predicting therapeutic outcome. The ability to monitor gene transfer, non-invasively, would strengthen the efficiency of therapy. In this regard, we have constructed and evaluated a replication-deficient adenovirus (Ad) containing the human somatostatin receptor subtype 2 (SSTR2) fused with a C-terminal yeast CD gene for the non-invasive monitoring of gene transfer and therapy. The resulting Ad (AdSSTR2-yCD) was evaluated in vitro in breast cancer cells to determine the function of the fusion protein. These studies demonstrated that the both the SSTR2 and yCD were functional in binding assays, conversion assays, and cytotoxicity assays. In vivo studies similarly demonstrated the functionality using conversion assays, biodistribution studies, and small animal positron-emission tomography (PET) imaging studies. In conclusion, the fusion protein has been validated as useful for the non-invasive imaging of yCD expression and will be evaluated in the future for monitoring yCD-based therapy. PMID:25837665

The ecology of insect-yeast relationships and its relevance to human industry.

PubMed

Madden, Anne A; Epps, Mary Jane; Fukami, Tadashi; Irwin, Rebecca E; Sheppard, John; Sorger, D Magdalena; Dunn, Robert R

2018-03-28

Many species of yeast are integral to human society. They produce many of our foods, beverages and industrial chemicals, challenge us as pathogens, and provide models for the study of our own biology. However, few species are regularly studied and much of their ecology remains unclear, hindering the development of knowledge that is needed to improve the relationships between humans and yeasts. There is increasing evidence that insects are an essential component of ascomycetous yeast ecology. We propose a 'dispersal-encounter hypothesis' whereby yeasts are dispersed by insects between ephemeral, spatially disparate sugar resources, and insects, in turn, obtain the benefits of an honest signal from yeasts for the sugar resources. We review the relationship between yeasts and insects through three main examples: social wasps, social bees and beetles, with some additional examples from fruit flies. Ultimately, we suggest that over the next decades, consideration of these ecological and evolutionary relationships between insects and yeasts will allow prediction of where new yeast diversity is most likely to be discovered, particularly yeasts with traits of interest to human industry. © 2018 The Author(s).

Assessment of yeast Saccharomyces cerevisiae component binding to Mycobacterium avium subspecies paratuberculosis using bovine epithelial cells.

PubMed

Li, Ziwei; You, Qiumei; Ossa, Faisury; Mead, Philip; Quinton, Margaret; Karrow, Niel A

2016-03-01

Since yeast Saccharomyces cerevisiae and its components are being used for the prevention and treatment of enteric diseases in different species, they may also be useful for preventing Johne's disease, a chronic inflammatory bowel disease of ruminants caused by Mycobacterium avium spp. paratuberculosis (MAP). This study aimed to identify potential yeast derivatives that may be used to help prevent MAP infection. The adherence of mCherry-labeled MAP to bovine mammary epithelial cell line (MAC-T cells) and bovine primary epithelial cells (BECs) co-cultured with yeast cell wall components (CWCs) from four different yeast strains (A, B, C and D) and two forms of dead yeast from strain A was investigated. The CWCs from all four yeast strains and the other two forms of dead yeast from strain A reduced MAP adhesion to MAC-T cells and BECs in a concentration-dependent manner after 6-h of exposure, with the dead yeast having the greatest effect. The following in vitro binding studies suggest that dead yeast and its' CWCs may be useful for reducing risk of MAP infection.

The ecology of insect–yeast relationships and its relevance to human industry

PubMed Central

Epps, Mary Jane; Sheppard, John; Sorger, D. Magdalena; Dunn, Robert R.

2018-01-01

Many species of yeast are integral to human society. They produce many of our foods, beverages and industrial chemicals, challenge us as pathogens, and provide models for the study of our own biology. However, few species are regularly studied and much of their ecology remains unclear, hindering the development of knowledge that is needed to improve the relationships between humans and yeasts. There is increasing evidence that insects are an essential component of ascomycetous yeast ecology. We propose a ‘dispersal–encounter hypothesis' whereby yeasts are dispersed by insects between ephemeral, spatially disparate sugar resources, and insects, in turn, obtain the benefits of an honest signal from yeasts for the sugar resources. We review the relationship between yeasts and insects through three main examples: social wasps, social bees and beetles, with some additional examples from fruit flies. Ultimately, we suggest that over the next decades, consideration of these ecological and evolutionary relationships between insects and yeasts will allow prediction of where new yeast diversity is most likely to be discovered, particularly yeasts with traits of interest to human industry. PMID:29563264

Genetic Polymorphism in Wine Yeasts: Mechanisms and Methods for Its Detection

PubMed Central

Guillamón, José M.; Barrio, Eladio

2017-01-01

The processes of yeast selection for using as wine fermentation starters have revealed a great phenotypic diversity both at interspecific and intraspecific level, which is explained by a corresponding genetic variation among different yeast isolates. Thus, the mechanisms involved in promoting these genetic changes are the main engine generating yeast biodiversity. Currently, an important task to understand biodiversity, population structure and evolutionary history of wine yeasts is the study of the molecular mechanisms involved in yeast adaptation to wine fermentation, and on remodeling the genomic features of wine yeast, unconsciously selected since the advent of winemaking. Moreover, the availability of rapid and simple molecular techniques that show genetic polymorphisms at species and strain levels have enabled the study of yeast diversity during wine fermentation. This review will summarize the mechanisms involved in generating genetic polymorphisms in yeasts, the molecular methods used to unveil genetic variation, and the utility of these polymorphisms to differentiate strains, populations, and species in order to infer the evolutionary history and the adaptive evolution of wine yeasts, and to identify their influence on their biotechnological and sensorial properties. PMID:28522998

Yeast Based Sensors

NASA Astrophysics Data System (ADS)

Shimomura-Shimizu, Mifumi; Karube, Isao

Since the first microbial cell sensor was studied by Karube et al. in 1977, many types of yeast based sensors have been developed as analytical tools. Yeasts are known as facultative anaerobes. Facultative anaerobes can survive in both aerobic and anaerobic conditions. The yeast based sensor consisted of a DO electrode and an immobilized omnivorous yeast. In yeast based sensor development, many kinds of yeast have been employed by applying their characteristics to adapt to the analyte. For example, Trichosporon cutaneum was used to estimate organic pollution in industrial wastewater. Yeast based sensors are suitable for online control of biochemical processes and for environmental monitoring. In this review, principles and applications of yeast based sensors are summarized.

The wine and beer yeast Dekkera bruxellensis

PubMed Central

Schifferdecker, Anna Judith; Dashko, Sofia; Ishchuk, Olena P; Piškur, Jure

2014-01-01

Recently, the non-conventional yeast Dekkera bruxellensis has been gaining more and more attention in the food industry and academic research. This yeast species is a distant relative of Saccharomyces cerevisiae and is especially known for two important characteristics: on the one hand, it is considered to be one of the main spoilage organisms in the wine and bioethanol industry; on the other hand, it is 'indispensable' as a contributor to the flavour profile of Belgium lambic and gueuze beers. Additionally, it adds to the characteristic aromatic properties of some red wines. Recently this yeast has also become a model for the study of yeast evolution. In this review we focus on the recently developed molecular and genetic tools, such as complete genome sequencing and transformation, to study and manipulate this yeast. We also focus on the areas that are particularly well explored in this yeast, such as the synthesis of off-flavours, yeast detection methods, carbon metabolism and evolutionary history. © 2014 The Authors. Yeast published by John Wiley & Sons, Ltd. PMID:24932634

The wine and beer yeast Dekkera bruxellensis.

PubMed

Schifferdecker, Anna Judith; Dashko, Sofia; Ishchuk, Olena P; Piškur, Jure

2014-09-01

Recently, the non-conventional yeast Dekkera bruxellensis has been gaining more and more attention in the food industry and academic research. This yeast species is a distant relative of Saccharomyces cerevisiae and is especially known for two important characteristics: on the one hand, it is considered to be one of the main spoilage organisms in the wine and bioethanol industry; on the other hand, it is 'indispensable' as a contributor to the flavour profile of Belgium lambic and gueuze beers. Additionally, it adds to the characteristic aromatic properties of some red wines. Recently this yeast has also become a model for the study of yeast evolution. In this review we focus on the recently developed molecular and genetic tools, such as complete genome sequencing and transformation, to study and manipulate this yeast. We also focus on the areas that are particularly well explored in this yeast, such as the synthesis of off-flavours, yeast detection methods, carbon metabolism and evolutionary history. © 2014 The Authors. Yeast published by John Wiley & Sons, Ltd.

Isolation and characterization of ethanol tolerant yeast strains

PubMed Central

Tikka, Chiranjeevi; Osuru, Hari Prasad; Atluri, Navya; Raghavulu, Praveen Chakravarthi Veera; yellapu, Nanda Kumar; Mannur, Ismail Shaik; Prasad, Uppu Venkateswara; Aluru, Sudheer; K, Narasimha Varma; Bhaskar, Matcha

2013-01-01

Yeast strains are commonly associated with sugar rich environments. Various fruit samples were selected as source for isolating yeast cells. The isolated cultures were identified at Genus level by colony morphology, biochemical characteristics and cell morphological characters. An attempt has been made to check the viability of yeast cells under different concentrations of ethanol. Ethanol tolerance of each strain was studied by allowing the yeast to grow in liquid YEPD (Yeast Extract Peptone Dextrose) medium having different concentrations of ethanol. A total of fifteen yeast strains isolated from different samples were used for the study. Seven strains of Saccharomyces cerevisiae obtained from different fruit sources were screened for ethanol tolerance. The results obtained in this study show a range of tolerance levels between 7%-12% in all the stains. Further, the cluster analysis based on 22 RAPD (Random Amplified polymorphic DNA) bands revealed polymorphisms in these seven Saccharomyces strains. PMID:23750092

Study of amyloids using yeast

PubMed Central

Wickner, Reed B.; Kryndushkin, Dmitry; Shewmaker, Frank; McGlinchey, Ryan; Edskes, Herman K.

2012-01-01

Summary Saccharomyces cerevisiae has been a useful model organism in such fields as the cell cycle, regulation of transcription, protein trafficking and cell biology, primarily because of its ease of genetic manipulation. This is no less so in the area of amyloid studies. The endogenous yeast amyloids described to date include prions, infectious proteins (Table 1), and some cell wall proteins (1). and amyloids of humans and a fungal prion have also been studied using the yeast system. Accordingly, the emphasis of this chapter will be on genetic, biochemical, cell biological and physical methods particularly useful in the study of yeast prions and other amyloids studied in yeast. We limit our description of these methods to those aspects which have been most useful in studying yeast prions, citing more detailed expositions in the literature. Volumes on yeast genetics methods (2–4), and on amyloids and prions (5, 6) are useful, and Masison has edited a volume of Methods on “Identification, analysis and characterization of fungal prions” which covers some of this territory (7). We also outline some useful physical methods, pointing the reader to more extensive and authoratative descriptions. PMID:22528100

Comparison of fermentative capacities of industrial baking and wild-type yeasts of the species Saccharomyces cerevisiae in different sugar media.

PubMed

Bell, P J; Higgins, V J; Attfield, P V

2001-04-01

To compare the fermentative capacity of wild and domesticated isolates of the genus Saccharomyces. The fermentative capacity of yeasts from a variety of wild and domesticated sources was tested in synthetic dough media that mimic major bread dough types. Domesticated yeast strains were found to have better maltose-utilizing capacity than wild yeast strains. The capacity to ferment sugars under high osmotic stress was randomly distributed amongst wild and baking strains of Saccharomyces. The domestication of bakers' yeast has enhanced the ability of yeasts to ferment maltose, without a similar impact on the fermentative capacity under high osmotic conditions. This study, combined with molecular studies of both wild and domesticated yeast, showed that domestication of bakers' yeast has resulted in improved maltose utilization, apparently via the duplication and mutation of the MAL genes.

Between science and industry-applied yeast research.

PubMed

Korhola, Matti

2018-03-01

I was fortunate to enter yeast research at the Alko Research Laboratories with a strong tradition in yeast biochemistry and physiology studies. At the same time in the 1980s there was a fundamental or paradigm change in molecular biology research with discoveries in DNA sequencing and other analytical and physical techniques for studying macromolecules and cells. Since that time biotechnological research has expanded the traditional fermentation industries to efficient production of industrial and other enzymes and specialty chemicals. Our efforts were directed towards improving the industrial production organisms: minerals enriched yeasts (Se, Cr, Zn) and high glutathione content yeast, baker´s, distiller´s, sour dough and wine yeasts, and the fungal Trichoderma reesei platform for enzyme production. I am grateful for the trust of my colleagues in several leadership positions at the Alko Research Laboratories, Yeast Industry Platform and at the international yeast community.

The canister around the FUSE satellite is removed on the pad at CCAS.

NASA Technical Reports Server (NTRS)

1999-01-01

At Launch Pad 17A, Cape Canaveral Air Station (CCAS), workers check out the protective cover placed over the top of NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite. The satellite is scheduled to be launched from CCAS June 23 aboard a Boeing Delta II rocket. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe.

Molecular mechanisms of autophagy in the cardiovascular system.

PubMed

Gatica, Damián; Chiong, Mario; Lavandero, Sergio; Klionsky, Daniel J

2015-01-30

Autophagy is a catabolic recycling pathway triggered by various intra- or extracellular stimuli that is conserved from yeast to mammals. During autophagy, diverse cytosolic constituents are enveloped by double-membrane vesicles, autophagosomes, which later fuse with lysosomes or the vacuole to degrade their cargo. Dysregulation in autophagy is associated with a diverse range of pathologies including cardiovascular disease, the leading cause of death in the world. As such, there is great interest in identifying novel mechanisms that govern the cardiovascular response to disease-related stress. First described in failing hearts, autophagy within the cardiovascular system has been characterized widely in cardiomyocytes, cardiac fibroblasts, endothelial cells, and vascular smooth muscle cells. In all cases, a window of optimal autophagic activity seems to be critical to the maintenance of cardiovascular homeostasis and function; excessive or insufficient levels of autophagic flux can each contribute to heart disease pathogenesis. Here, we review the molecular mechanisms that govern autophagosome formation and analyze the link between autophagy and cardiovascular disease. © 2015 American Heart Association, Inc.

A family of metal-dependent phosphatases implicated in metabolite damage-control

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Lili; Khusnutdinova, Anna; Nocek, Boguslaw

DUF89 family proteins occur widely in both prokaryotes and eukaryotes, but their functions are unknown. Here we define three DUF89 subfamilies (I, II, and III), with subfamily II being split into stand-alone proteins and proteins fused to pantothenate kinase (PanK). We demonstrated that DUF89 proteins have metal-dependent phosphatase activity against reactive phosphoesters or their damaged forms, notably sugar phosphates (subfamilies II and III), phosphopantetheine and its S-sulfonate or sulfonate (subfamily II-PanK fusions), and nucleotides (subfamily I). Genetic and comparative genomic data strongly associated DUF89 genes with phosphoester metabolism. The crystal structure of the yeast (Saccharomyces cerevisiae) subfamily III protein YMR027Wmore » revealed a novel phosphatase active site with fructose 6-phosphate and Mg2+ bound near conserved signature residues Asp254 and Asn255 that are critical for activity. These findings indicate that DUF89 proteins are previously unrecognized hydrolases whose characteristic in vivo function is to limit potentially harmful buildups of normal or damaged phosphometabolites.« less

A family of metal-dependent phosphatases implicated in metabolite damage-control

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Lili; Shanklin, John; Khusnutdinova, Anna

DUF89 family proteins occur widely in pro- and eukaryotes but their functions are unknown. Here we define three DUF89 subfamilies (I, II, and III), subfamily II being split into standalone proteins and proteins fused to pantothenate kinase (PanK). We demonstrated that DUF89 proteins have metaldependent phosphatase activity against reactive phosphoesters or their damaged forms, notably sugar phosphates (subfamilies II and III), phosphopantetheine and its S-sulfonate or sulfonate (subfamily II-PanK fusions), and nucleotides (subfamily I). Genetic and comparative genomic data strongly associated DUF89 genes with phosphoester metabolism. The crystal structure of the yeast (Saccharomyces cerevisiae) subfamily III protein YMR027W revealed amore » novel phosphatase active site with fructose 6-phosphate and Mg 2+ bound near conserved signature residues Asp254 and Asn255 that are critical for activity. These findings indicate that DUF89 proteins are previously unrecognized hydrolases whose characteristic in vivo function is to limit potentially harmful buildups of normal or damaged phosphometabolites.« less

A single-chain TALEN architecture for genome engineering.

PubMed

Sun, Ning; Zhao, Huimin

2014-03-04

Transcription-activator like effector nucleases (TALENs) are tailor-made DNA endonucleases and serve as a powerful tool for genome engineering. Site-specific DNA cleavage can be made by the dimerization of FokI nuclease domains at custom-targeted genomic loci, where a pair of TALENs must be positioned in close proximity with an appropriate orientation. However, the simultaneous delivery and coordinated expression of two bulky TALEN monomers (>100 kDa) in cells may be problematic to implement for certain applications. Here, we report the development of a single-chain TALEN (scTALEN) architecture, in which two FokI nuclease domains are fused on a single polypeptide. The scTALEN was created by connecting two FokI nuclease domains with a 95 amino acid polypeptide linker, which was isolated from a linker library by high-throughput screening. We demonstrated that scTALENs were catalytically active as monomers in yeast and human cells. The use of this novel scTALEN architecture should reduce protein payload, simplify design and decrease production cost.

Characterization of a Trifunctional Mimivirus mRNA Capping Enzyme and Crystal Structure of the RNA Triphosphatase Domain

DOE Office of Scientific and Technical Information (OSTI.GOV)

Benarroch,D.; Smith, P.; Shuman, S.

2008-01-01

The RNA triphosphatase (RTPase) components of the mRNA capping apparatus are a bellwether of eukaryal taxonomy. Fungal and protozoal RTPases belong to the triphosphate tunnel metalloenzyme (TTM) family, exemplified by yeast Cet1. Several large DNA viruses encode metal-dependent RTPases unrelated to the cysteinyl-phosphatase RTPases of their metazoan host organisms. The origins of DNA virus RTPases are unclear because they are structurally uncharacterized. Mimivirus, a giant virus of amoeba, resembles poxviruses in having a trifunctional capping enzyme composed of a metal-dependent RTPase module fused to guanylyltransferase (GTase) and guanine-N7 methyltransferase domains. The crystal structure of mimivirus RTPase reveals a minimized tunnelmore » fold and an active site strikingly similar to that of Cet1. Unlike homodimeric fungal RTPases, mimivirus RTPase is a monomer. The mimivirus TTM-type RTPase-GTase fusion resembles the capping enzymes of amoebae, providing evidence that the ancestral large DNA virus acquired its capping enzyme from a unicellular host.« less

The Zinc-Finger Thylakoid-Membrane Protein FIP Is Involved With Abiotic Stress Response in Arabidopsis thaliana.

PubMed

Lopes, Karina L; Rodrigues, Ricardo A O; Silva, Marcos C; Braga, Wiliane G S; Silva-Filho, Marcio C

2018-01-01

Many plant genes have their expression modulated by stress conditions. Here, we used Arabidopsis FtsH5 protease, which expression is regulated by light stress, as bait in a yeast two-hybrid screen to search for new proteins involved in the stress response. As a result, we found FIP (FtsH5 Interacting Protein), which possesses an amino proximal cleavable transit peptide, a hydrophobic membrane-anchoring region, and a carboxyl proximal C 4 -type zinc-finger domain. In vivo experiments using FIP fused to green fluorescent protein (GFP) showed a plastid localization. This finding was corroborated by chloroplast import assays that showed FIP inserted in the thylakoid membrane. FIP expression was down-regulated in plants exposed to high light intensity, oxidative, salt, and osmotic stresses, whereas mutant plants expressing low levels of FIP were more tolerant to these abiotic stresses. Our data shows a new thylakoid-membrane protein involved with abiotic stress response in Arabidopsis thaliana .

Using constitutive activity to define appropriate high-throughput screening assays for orphan g protein-coupled receptors.

PubMed

Ngo, Tony; Coleman, James L J; Smith, Nicola J

2015-01-01

Orphan G protein-coupled receptors represent an underexploited resource for drug discovery but pose a considerable challenge for assay development because their cognate G protein signaling pathways are often unknown. In this methodological chapter, we describe the use of constitutive activity, that is, the inherent ability of receptors to couple to their cognate G proteins in the absence of ligand, to inform the development of high-throughput screening assays for a particular orphan receptor. We specifically focus on a two-step process, whereby constitutive G protein coupling is first determined using yeast Gpa1/human G protein chimeras linked to growth and β-galactosidase generation. Coupling selectivity is then confirmed in mammalian cells expressing endogenous G proteins and driving accumulation of transcription factor-fused luciferase reporters specific to each of the classes of G protein. Based on these findings, high-throughput screening campaigns can be performed on the already miniaturized mammalian reporter system.

An auto-biotinylated bifunctional protein nanowire for ultra-sensitive molecular biosensing.

PubMed

Men, Dong; Zhang, Zhi-Ping; Guo, Yong-Chao; Zhu, Duan-Hao; Bi, Li-Jun; Deng, Jiao-Yu; Cui, Zong-Qiang; Wei, Hong-Ping; Zhang, Xian-En

2010-12-15

In order to obtain an ultra-sensitive molecular biosensor, we designed an auto-biotinylated bifunctional protein nanowire (bFPNw) based on the self-assembly of a yeast amyloid protein, Sup35, to which protein G and a biotin acceptor peptide (BAP) were genetically fused. These auto-biotinylated bFPNws can transfer hundreds of commercially available diagnostic enzymes to an antigen-antibody complex via the biotin-avidin system, greatly enhancing the sensitivity of immune-biosensing. Compared to our previously reported seeding-induced bFPNws (Men et al., 2009), these auto-biotinylated bFPNws gave greater signal amplification, reduced non-specific binding and improved stability. The auto-biotinylated self-assembled bFPNw molecular biosensors were applied to detect Yersinia pestis (Y. pestis) F1 antigen and showed a 2000- to 4000-fold increase in sensitivity compared to traditional immunoassays, demonstrating the potential use of these self-assembling protein nanowires in biosensing. Copyright © 2010 Elsevier B.V. All rights reserved.

Tunable and reversible drug control of protein production via a self-excising degron.

PubMed

Chung, Hokyung K; Jacobs, Conor L; Huo, Yunwen; Yang, Jin; Krumm, Stefanie A; Plemper, Richard K; Tsien, Roger Y; Lin, Michael Z

2015-09-01

An effective method for direct chemical control over the production of specific proteins would be widely useful. We describe small molecule-assisted shutoff (SMASh), a technique in which proteins are fused to a degron that removes itself in the absence of drug, resulting in the production of an untagged protein. Clinically tested HCV protease inhibitors can then block degron removal, inducing rapid degradation of subsequently synthesized copies of the protein. SMASh allows reversible and dose-dependent shutoff of various proteins in multiple mammalian cell types and in yeast. We also used SMASh to confer drug responsiveness onto an RNA virus for which no licensed inhibitors exist. As SMASh does not require the permanent fusion of a large domain, it should be useful when control over protein production with minimal structural modification is desired. Furthermore, as SMASh involves only a single genetic modification and does not rely on modulating protein-protein interactions, it should be easy to generalize to multiple biological contexts.

A family of metal-dependent phosphatases implicated in metabolite damage-control

DOE PAGES

Huang, Lili; Shanklin, John; Khusnutdinova, Anna; ...

2016-06-20

DUF89 family proteins occur widely in pro- and eukaryotes but their functions are unknown. Here we define three DUF89 subfamilies (I, II, and III), subfamily II being split into standalone proteins and proteins fused to pantothenate kinase (PanK). We demonstrated that DUF89 proteins have metaldependent phosphatase activity against reactive phosphoesters or their damaged forms, notably sugar phosphates (subfamilies II and III), phosphopantetheine and its S-sulfonate or sulfonate (subfamily II-PanK fusions), and nucleotides (subfamily I). Genetic and comparative genomic data strongly associated DUF89 genes with phosphoester metabolism. The crystal structure of the yeast (Saccharomyces cerevisiae) subfamily III protein YMR027W revealed amore » novel phosphatase active site with fructose 6-phosphate and Mg 2+ bound near conserved signature residues Asp254 and Asn255 that are critical for activity. These findings indicate that DUF89 proteins are previously unrecognized hydrolases whose characteristic in vivo function is to limit potentially harmful buildups of normal or damaged phosphometabolites.« less

Yeasts in floral nectar: a quantitative survey

PubMed Central

Herrera, Carlos M.; de Vega, Clara; Canto, Azucena; Pozo, María I.

2009-01-01

Background and Aims One peculiarity of floral nectar that remains relatively unexplored from an ecological perspective is its role as a natural habitat for micro-organisms. This study assesses the frequency of occurrence and abundance of yeast cells in floral nectar of insect-pollinated plants from three contrasting plant communities on two continents. Possible correlations between interspecific differences in yeast incidence and pollinator composition are also explored. Methods The study was conducted at three widely separated areas, two in the Iberian Peninsula (Spain) and one in the Yucatán Peninsula (Mexico). Floral nectar samples from 130 species (37–63 species per region) in 44 families were examined microscopically for the presence of yeast cells. For one of the Spanish sites, the relationship across species between incidence of yeasts in nectar and the proportion of flowers visited by each of five major pollinator categories was also investigated. Key Results Yeasts occurred regularly in the floral nectar of many species, where they sometimes reached extraordinary densities (up to 4 × 105 cells mm−3). Depending on the region, between 32 and 44 % of all nectar samples contained yeasts. Yeast cell densities in the order of 104 cells mm−3 were commonplace, and densities >105 cells mm−3 were not rare. About one-fifth of species at each site had mean yeast cell densities >104 cells mm−3. Across species, yeast frequency and abundance were directly correlated with the proportion of floral visits by bumble-bees, and inversely with the proportion of visits by solitary bees. Conclusions Incorporating nectar yeasts into the scenario of plant–pollinator interactions opens up a number of intriguing avenues for research. In addition, with yeasts being as ubiquitous and abundant in floral nectars as revealed by this study, and given their astounding metabolic versatility, studies focusing on nectar chemical features should carefully control for the presence of yeasts in nectar samples. PMID:19208669

Clinical and tree hollow populations of human pathogenic yeast in Hamilton, Ontario, Canada are different.

PubMed

Carvalho, Chris; Yang, Jiaqi; Vogan, Aaron; Maganti, Harinad; Yamamura, Deborah; Xu, Jianping

2014-05-01

Yeast are among the most frequent pathogens in humans. The dominant yeast causing human infections belong to the genus Candida and Candida albicans is the most frequently isolated species. However, several non-C. albicans species are becoming increasingly common in patients worldwide. The relationships between yeast in humans and the natural environments remain poorly understood. Furthermore, it is often difficult to identify or exclude the origins of disease-causing yeast from specific environmental reservoirs. In this study, we compared the yeast isolates from tree hollows and from clinics in Hamilton, Ontario, Canada. Our surveys and analyses showed significant differences in yeast species composition, in their temporal dynamics, and in yeast genotypes between isolates from tree hollows and hospitals. Our results are inconsistent with the hypothesis that yeast from trees constitute a significant source of pathogenic yeast in humans in this region. Similarly, the yeast in humans and clinics do not appear to contribute to yeast in tree hollows. © 2013 Blackwell Verlag GmbH.

Nectar-living yeasts of a tropical host plant community: diversity and effects on community-wide floral nectar traits

PubMed Central

2017-01-01

We characterize the diversity of nectar-living yeasts of a tropical host plant community at different hierarchical sampling levels, measure the associations between yeasts and nectariferous plants, and measure the effect of yeasts on nectar traits. Using a series of hierarchically nested sampling units, we extracted nectar from an assemblage of host plants that were representative of the diversity of life forms, flower shapes, and pollinator types in the tropical area of Yucatan, Mexico. Yeasts were isolated from single nectar samples; their DNA was identified, the yeast cell density was estimated, and the sugar composition and concentration of nectar were quantified using HPLC. In contrast to previous studies from temperate regions, the diversity of nectar-living yeasts in the plant community was characterized by a relatively high number of equally common species with low dominance. Analyses predict highly diverse nectar yeast communities in a relatively narrow range of tropical vegetation, suggesting that the diversity of yeasts will increase as the number of sampling units increases at the level of the species, genera, and botanical families of the hosts. Significant associations between specific yeast species and host plants were also detected; the interaction between yeasts and host plants impacted the effect of yeast cell density on nectar sugars. This study provides an overall picture of the diversity of nectar-living yeasts in tropical host plants and suggests that the key factor that affects the community-wide patterns of nectar traits is not nectar chemistry, but rather the type of yeasts interacting with host plants. PMID:28717591

Assessment of Multi Fragment Melting Analysis System (MFMAS) for the Identification of Food-Borne Yeasts.

PubMed

Kesmen, Zülal; Büyükkiraz, Mine E; Özbekar, Esra; Çelik, Mete; Özkök, F Özge; Kılıç, Özge; Çetin, Bülent; Yetim, Hasan

2018-06-01

Multi Fragment Melting Analysis System (MFMAS) is a novel approach that was developed for the species-level identification of microorganisms. It is a software-assisted system that performs concurrent melting analysis of 8 different DNA fragments to obtain a fingerprint of each strain analyzed. The identification is performed according to the comparison of these fingerprints with the fingerprints of known yeast species recorded in a database to obtain the best possible match. In this study, applicability of the yeast version of the MFMAS (MFMAS-yeast) was evaluated for the identification of food-associated yeast species. For this purpose, in this study, a total of 145 yeast strains originated from foods and beverages and 19 standard yeast strains were tested. The DNAs isolated from these yeast strains were analyzed by the MFMAS, and their species were successfully identified with a similarity rate of 95% or higher. It was shown that the strains belonged to 43 different yeast species that are widely found in the foods. A clear discrimination was also observed in the phylogenetically related species. In conclusion, it might be suggested that the MFMAS-yeast seems to be a highly promising approach for a rapid, accurate, and one-step identification of the yeasts isolated from food products and/or their processing environments.

29 CFR 1910.399 - Definitions applicable to this subpart.

Code of Federal Regulations, 2010 CFR

2010-07-01

... assembly of a fuse support with either a fuseholder, fuse carrier, or disconnecting blade. The fuseholder or fuse carrier may include a conducting element (fuse link), or may act as the disconnecting blade... in which all or part of the fuse support and its fuse link or disconnecting blade are mounted in oil...

29 CFR 1910.399 - Definitions applicable to this subpart.

Code of Federal Regulations, 2013 CFR

2013-07-01

... assembly of a fuse support with either a fuseholder, fuse carrier, or disconnecting blade. The fuseholder or fuse carrier may include a conducting element (fuse link), or may act as the disconnecting blade... in which all or part of the fuse support and its fuse link or disconnecting blade are mounted in oil...

29 CFR 1910.399 - Definitions applicable to this subpart.

Code of Federal Regulations, 2014 CFR

2014-07-01

... assembly of a fuse support with either a fuseholder, fuse carrier, or disconnecting blade. The fuseholder or fuse carrier may include a conducting element (fuse link), or may act as the disconnecting blade... in which all or part of the fuse support and its fuse link or disconnecting blade are mounted in oil...

29 CFR 1910.399 - Definitions applicable to this subpart.

Code of Federal Regulations, 2011 CFR

2011-07-01

... assembly of a fuse support with either a fuseholder, fuse carrier, or disconnecting blade. The fuseholder or fuse carrier may include a conducting element (fuse link), or may act as the disconnecting blade... in which all or part of the fuse support and its fuse link or disconnecting blade are mounted in oil...

29 CFR 1910.399 - Definitions applicable to this subpart.

Code of Federal Regulations, 2012 CFR

2012-07-01

... assembly of a fuse support with either a fuseholder, fuse carrier, or disconnecting blade. The fuseholder or fuse carrier may include a conducting element (fuse link), or may act as the disconnecting blade... in which all or part of the fuse support and its fuse link or disconnecting blade are mounted in oil...

Genotoxicity Assessment of Drinking Water Disinfection Byproducts by DNA Damage and Repair Pathway Profiling Analysis.

PubMed

Lan, Jiaqi; Rahman, Sheikh Mokhlesur; Gou, Na; Jiang, Tao; Plewa, Micheal J; Alshawabkeh, Akram; Gu, April Z

2018-06-05

Genotoxicity is considered a major concern for drinking water disinfection byproducts (DBPs). Of over 700 DBPs identified to date, only a small number has been assessed with limited information for DBP genotoxicity mechanism(s). In this study, we evaluated genotoxicity of 20 regulated and unregulated DBPs applying a quantitative toxicogenomics approach. We used GFP-fused yeast strains that examine protein expression profiling of 38 proteins indicative of all known DNA damage and repair pathways. The toxicogenomics assay detected genotoxicity potential of these DBPs that is consistent with conventional genotoxicity assays end points. Furthermore, the high-resolution, real-time pathway activation and protein expression profiling, in combination with clustering analysis, revealed molecular level details in the genotoxicity mechanisms among different DBPs and enabled classification of DBPs based on their distinct DNA damage effects and repair mechanisms. Oxidative DNA damage and base alkylation were confirmed to be the main molecular mechanisms of DBP genotoxicity. Initial exploration of QSAR modeling using moleular genotoxicity end points (PELI) suggested that genotoxicity of DBPs in this study was correlated with topological and quantum chemical descriptors. This study presents a toxicogenomics-based assay for fast and efficient mechanistic genotoxicity screening and assessment of a large number of DBPs. The results help to fill in the knowledge gap in the understanding of the molecular mechanisms of DBP genotoxicity.

Distortion of three-unit implant frameworks during casting, soldering, and simulated porcelain firings.

PubMed

Zervas, P J; Papazoglou, E; Beck, F M; Carr, A B

1999-09-01

The aim of this study was to assess distortion inherent in casting, soldering, and simulated porcelain firings of screw-retained, implant-supported three-unit fixed partial dentures (FPDs). Ten wax patterns were fabricated on a die-stone cast containing two implants, 20 mm apart from center to center. Five specimens were cast in a high-palladium alloy, exposed to simulated porcelain firings, sectioned, and then soldered with low-fusing solder. Five specimens were cast, sectioned, soldered with high-fusing solder, and then exposed to simulated porcelain firings. For each specimen, two horizontal and six vertical distances between appropriately scribed reference points were measured with a traveling microscope. Comparisons were made among the various measurements taken after wax-pattern fabrication, casting, high- and low-fusing soldering, and each porcelain firing. Data were analyzed using a repeated-measures factorial ANOVA (alpha = 0.05). Significant difference was detected in the amount of horizontal distortion during casting (53 +/- 24 microns) and high-fusing soldering (-49 +/- 50 microns), as well as in the amount of horizontal distortion during high-fusing soldering (-49 +/- 50 microns) and low-fusing soldering (17 +/- 26 microns). However, no clinically significant difference was found in the amount of horizontal distortion during casting, low-fusing, and high-fusing soldering. The greatest amount of distortion during the simulated porcelain firings took place during the oxidizing cycle. Soldering did not improve the casting misfit of a three-unit implant-retained FPD model. Metal-ceramic implant frameworks should be oxidized before intraoral fit evaluation.

Interactions between Brettanomyces bruxellensis and other yeast species during the initial stages of winemaking.

PubMed

Renouf, V; Falcou, M; Miot-Sertier, C; Perello, M C; De Revel, G; Lonvaud-Funel, A

2006-06-01

Wine is the product of complex interactions between yeasts and bacteria in grape must. Amongst yeast populations, two groups can be distinguished. The first, named non-Saccharomyces (NS), colonizes, with many other micro-organisms, the surface of grape berries. In the past, NS yeasts were primarily considered as spoilage micro-organisms. However, recent studies have established a positive contribution of certain NS yeasts to wine quality. Amongst the group of NS yeasts, Brettanomyces bruxellensis, which is not prevalent on wine grapes, plays an important part in the evolution of wine aroma. Some of their secondary metabolites, namely volatile phenols, are responsible for wine spoilage. The other group contributing to wine aroma, which is also the main agent of alcoholic fermentation (AF), is composed of Saccharomyces species. The fermenting must is a complex microbial ecosystem where numerous yeast strains grow and die according to their adaptation to the medium. Yeast-yeast interactions occur during winemaking right from the onset of AF. The aim of this study was to describe the interactions between B. bruxellensis, other NS and Saccharomyces cerevisiae during laboratory and practical scale winemaking. Molecular methods such as internal transcribed spacer-restriction fragment length polymorphism and polymerase chain reaction and denaturing gradient gel electrophoresis were used in laboratory scale experiments and cellar observations. The influence of different oenological practices, like the level of sulphiting at harvest time, cold maceration preceding AF, addition of commercial active dry yeasts on B. bruxellensis and other yeast interactions and their evolution during the initial stages of winemaking have been studied. Brettanomyces bruxellensis was the most adapted NS yeast at the beginning of AF, and towards the end of AF it appeared to be more resistant than S. cerevisiae to the conditions of increased alcohol and sugar limitation. Among all NS yeast species, B. bruxellensis is better adapted than other wild yeasts to resist in must and during AF. Moreover, B. bruxellensis appeared to be more tolerant to ethanol stress than S. cerevisiae and after AF B. bruxellensis was the main yeast species in wine. Brettanomyces bruxellensis interacts with other yeast species and adapts to the wine medium as the dominant yeast species at the end of AF. Contamination of B. bruxellensis might take place at the beginning of malolactic fermentation, which is a critical stage in winemaking.

A Mission Concept for FUSE Operations in GFY09 and Beyond

NASA Astrophysics Data System (ADS)

Moos, H. W.; Sonneborn, G.; Blair, W. P.; Kruk, J. W.; FUSE Science Operations Team

2007-05-01

We are developing a new mission concept for the Far Ultraviolet Spectroscopic Explorer (FUSE) that would significantly reduce operating costs but would continue the availability of this unique resource into GFY09 and beyond. Launched in 1999, the FUSE satellite obtains R=20,000 spectra of astronomical sources in the far-ultraviolet (912 - 1187 A) wavelength range. The FUSE scientific instrument remains healthy and the satellite has made a remarkable recovery from attitude control problems in late 2004. We expect FUSE to remain a viable scientific tool for the foreseeable future. Current plans for FUSE operations extend through GFY2008 (Sept. 30, 2008). Key elements of this new mission concept include a) continued automation and streamlining of operations to reduce costs, and b) an emphasis on a small number of unique, high priority science programs, particularly those requiring integration times on key targets that are significantly longer than has been possible in the mission design to date. A prime example of the latter would be 100 - 400 ks integrations on selected quasars to provide much improved diagnostic power to study the intergalactic medium. Synergy with the scientific objectives of the Cosmic Origins Spectrograph (COS) program on HST, and the complementary nature of FUSE and COS data on the same sightlines, is but one major motivation for this operations model. In addition to programs emphasizing very long integrations, opportunities for other high priority targets would exist. We will describe some of the ongoing development toward such an operations model as well as the scientific drivers discussed to date. Community input on these and other science drivers for extended FUSE operations is encouraged. FUSE is operated for NASA by Johns Hopkins University under NASA contract NAS5-32985.

Glycosylceramide modifies the flavor and metabolic characteristics of sake yeast.

PubMed

Ferdouse, Jannatul; Yamamoto, Yuki; Taguchi, Seiga; Yoshizaki, Yumiko; Takamine, Kazunori; Kitagaki, Hiroshi

2018-01-01

In the manufacture of sake, Japanese traditional rice wine, sake yeast is fermented with koji, which is steamed rice fermented with the non-pathogenic fungus Aspergillus oryzae . During fermentation, sake yeast requires lipids, such as unsaturated fatty acids and sterols, in addition to substances provided by koji enzymes for fermentation. However, the role of sphingolipids on the brewing characteristics of sake yeast has not been studied. In this study, we revealed that glycosylceramide, one of the sphingolipids abundant in koji, affects yeast fermentation. The addition of soy, A. oryzae , and Grifola frondosa glycosylceramide conferred a similar effect on the flavor profiles of sake yeast. In particular, the addition of A. oryzae and G. frondosa glycosylceramide were very similar in terms of the decreases in ethyl caprylate and ethyl 9-decenoate. The addition of soy glycosylceramide induced metabolic changes to sake yeast such as a decrease in glucose, increases in ethanol and glycerol and changes in several amino acids and organic acids concentrations. Tricarboxylic acid (TCA) cycle, pyruvate metabolism, starch and sucrose metabolism, and glycerolipid metabolism were overrepresented in the cultures incubated with sake yeast and soy glycosylceramide. This is the first study of the effect of glycosylceramide on the flavor and metabolic profile of sake yeast.

Characterization and Dynamic Behavior of Wild Yeast during Spontaneous Wine Fermentation in Steel Tanks and Amphorae

PubMed Central

Díaz, Cecilia; Molina, Ana María; Nähring, Jörg; Fischer, Rainer

2013-01-01

We studied the dynamic behavior of wild yeasts during spontaneous wine fermentation at a winery in the Valais region of Switzerland. Wild yeasts in the winery environment were characterized using a PCR-RFLP method. Up to 11 different yeast species were isolated from the vineyard air, whereas only seven were recovered from the grapes surface. We initially investigated a cultureindependent method in pilot-scale steel fermentation tanks and found a greater diversity of yeasts in the musts from two red grape varieties compared to three white grape varieties. We found that the yeasts Metschnikowia pulcherrima, Rhodotorula mucilaginosa, Pichia kluyveri, P. membranifaciens and Saccharomyces cerevisiae remained active at the end of the fermentation. We also studied the dynamic behavior of yeasts in Qvevris for the first time using a novel, highlysensitive quantitative real-time PCR method. We found that non-Saccharomyces yeasts were present during the entire fermentation process, with R. mucilaginosa and P. anomala the most prominent species. We studied the relationship between the predominance of different species and the output of the fermentation process. We identified so-called spoilage yeasts in all the fermentations, but high levels of acetic acid accumulated only in those fermentations with an extended lag phase. PMID:23738327

Glycosylceramide modifies the flavor and metabolic characteristics of sake yeast

PubMed Central

Taguchi, Seiga; Yoshizaki, Yumiko; Takamine, Kazunori

2018-01-01

In the manufacture of sake, Japanese traditional rice wine, sake yeast is fermented with koji, which is steamed rice fermented with the non-pathogenic fungus Aspergillus oryzae. During fermentation, sake yeast requires lipids, such as unsaturated fatty acids and sterols, in addition to substances provided by koji enzymes for fermentation. However, the role of sphingolipids on the brewing characteristics of sake yeast has not been studied. In this study, we revealed that glycosylceramide, one of the sphingolipids abundant in koji, affects yeast fermentation. The addition of soy, A. oryzae, and Grifola frondosa glycosylceramide conferred a similar effect on the flavor profiles of sake yeast. In particular, the addition of A. oryzae and G. frondosa glycosylceramide were very similar in terms of the decreases in ethyl caprylate and ethyl 9-decenoate. The addition of soy glycosylceramide induced metabolic changes to sake yeast such as a decrease in glucose, increases in ethanol and glycerol and changes in several amino acids and organic acids concentrations. Tricarboxylic acid (TCA) cycle, pyruvate metabolism, starch and sucrose metabolism, and glycerolipid metabolism were overrepresented in the cultures incubated with sake yeast and soy glycosylceramide. This is the first study of the effect of glycosylceramide on the flavor and metabolic profile of sake yeast. PMID:29761062

HVI Ballistic Limit Characterization of Fused Silica Thermal Panes

NASA Technical Reports Server (NTRS)

Miller, J. E.; Bohl, W. D.; Christiansen, E. L.; Davis, B. A.; Deighton, K. D.

2015-01-01

Fused silica window systems are used heavily on crewed reentry vehicles, and they are currently being used on the next generation of US crewed spacecraft, Orion. These systems improve crew situational awareness and comfort, as well as, insulating the reentry critical components of a spacecraft against the intense thermal environments of atmospheric reentry. Additionally, these materials are highly exposed to space environment hazards like solid particle impacts. This paper discusses impact studies up to 10 km/s on a fused silica window system proposed for the Orion spacecraft. A ballistic limit equation that describes the threshold of perforation of a fuse silica pane over a broad range of impact velocities, obliquities and projectile materials is discussed here.

Manipulation of culture conditions alters lipid content and fatty acid profiles of a wide variety of known and new oleaginous yeast species.

PubMed

Sitepu, Irnayuli R; Sestric, Ryan; Ignatia, Laura; Levin, David; German, J Bruce; Gillies, Laura A; Almada, Luis A G; Boundy-Mills, Kyria L

2013-09-01

Oleaginous yeasts have been studied for oleochemical production for over 80 years. Only a few species have been studied intensely. To expand the diversity of oleaginous yeasts available for lipid research, we surveyed a broad diversity of yeasts with indicators of oleaginicity including known oleaginous clades, and buoyancy. Sixty-nine strains representing 17 genera and 50 species were screened for lipid production. Yeasts belonged to Ascomycota families, Basidiomycota orders, and the yeast-like algal genus Prototheca. Total intracellular lipids and fatty acid composition were determined under different incubation times and nitrogen availability. Thirteen new oleaginous yeast species were discovered, representing multiple ascomycete and basidiomycete clades. Nitrogen starvation generally increased intracellular lipid content. The fatty acid profiles varied with the growth conditions regardless of taxonomic affiliation. The dominant fatty acids were oleic acid, palmitic acid, linoleic acid, and stearic acid. Yeasts and culture conditions that produced fatty acids appropriate for biodiesel were identified. Copyright © 2013 Elsevier Ltd. All rights reserved.

Manipulation of culture conditions alters lipid content and fatty acid profiles of a wide variety of known and new oleaginous yeasts species

PubMed Central

Sitepu, Irnayuli R.; Sestric, Ryan; Ignatia, Laura; Levin, David; German, J. Bruce; Gillies, Laura A.; Almada, Luis A.G.; Boundy-Mills, Kyria L.

2013-01-01

Oleaginous yeasts have been studied for oleochemical production for over 80 years. Only a few species have been studied intensely. To expand the diversity of oleaginous yeasts available for lipid research, we surveyed a broad diversity of yeasts with indicators of oleaginicity including known oleaginous clades, and buoyancy. Sixty-nine strains representing 17 genera and 50 species were screened for lipid production. Yeasts belonged to Ascomycota families, Basidiomycota orders, and the yeast-like algal genus Prototheca. Total intracellular lipids and fatty acid composition were determined under different incubation times and nitrogen availability. Thirteen new oleaginous yeast species were discovered, representing multiple ascomycete and basidiomycete clades. Nitrogen starvation generally increased intracellular lipid content. The fatty acid profiles varied with the growth conditions regardless of taxonomic affiliation. The dominant fatty acids were oleic acid, palmitic acid, linoleic acid, and stearic acid. Yeasts and culture conditions that produced fatty acids appropriate for biodiesel were identified. PMID:23891835

Yeast Infection and Diabetes Mellitus among Pregnant Mother in Malaysia

PubMed Central

Sopian, Iylia Liyana; Shahabudin, Sa’adiah; Ahmed, Mowaffaq Adam; Lung, Leslie Than Thian; Sandai, Doblin

2016-01-01

Background Vaginal yeast infection refers to irritation of the vagina due to the presence of opportunistic yeast of the genus Candida (mostly Candida albicans). About 75% of women will have at least one episode of vaginal yeast infection during their lifetime. Several studies have shown that pregnancy and uncontrolled diabetes increase the infection risk. Reproductive hormone fluctuations during pregnancy and elevated glucose levels characteristic of diabetes provide the carbon needed for Candida overgrowth and infection. The goal of this study was to determine the prevalence of vaginal yeast infection among pregnant women with and without diabetes. Methods This was a case-control study using cases reports from Kepala Batas Health Clinic, Penang State, Malaysia from 2006 to 2012. In total, 740 pregnant ladies were chosen as sample of which 370 were diabetic and 370 were non-diabetic cases. Results No relationship between diabetes and the occurrence of vaginal yeast infection in pregnant women was detected, and there was no significant association between infection and age group, race or education level. Conclusion In conclusion, within radius of this study, vaginal yeast infection can occur randomly in pregnant women. PMID:27540323

An Ancient Yeast for Young Geneticists: A Primer on the Schizosaccharomyces pombe Model System

PubMed Central

Hoffman, Charles S.; Wood, Valerie; Fantes, Peter A.

2015-01-01

The fission yeast Schizosaccharomyces pombe is an important model organism for the study of eukaryotic molecular and cellular biology. Studies of S. pombe, together with studies of its distant cousin, Saccharomyces cerevisiae, have led to the discovery of genes involved in fundamental mechanisms of transcription, translation, DNA replication, cell cycle control, and signal transduction, to name but a few processes. However, since the divergence of the two species approximately 350 million years ago, S. pombe appears to have evolved less rapidly than S. cerevisiae so that it retains more characteristics of the common ancient yeast ancestor, causing it to share more features with metazoan cells. This Primer introduces S. pombe by describing the yeast itself, providing a brief description of the origins of fission yeast research, and illustrating some genetic and bioinformatics tools used to study protein function in fission yeast. In addition, a section on some key differences between S. pombe and S. cerevisiae is included for readers with some familiarity with budding yeast research but who may have an interest in developing research projects using S. pombe. PMID:26447128

Yeast communities in Sphagnum phyllosphere along the temperature-moisture ecocline in the boreal forest-swamp ecosystem and description of Candida sphagnicola sp. nov.

PubMed

Kachalkin, Aleksey V; Yurkov, Andrey M

2012-06-01

The effects of the temperature-moisture factors on the phylloplane yeast communities inhabiting Sphagnum mosses were studied along the transition from a boreal forest to a swamp biotope at the Central Forest State Biosphere Reserve (Tver region, Russia). We tested the hypothesis that microclimatic parameters affect yeast community composition and structure even on a rather small spatial scale. Using a conventional plating technique we isolated and identified by molecular methods a total of 15 species of yeasts. Total yeast counts and species richness values did not depend on environmental factors, although yeast community composition and structure did. On average, Sphagnum in the swamp biotope supported a more evenly structured yeast community. Relative abundance of ascomycetous yeasts was significantly higher on swamp moss. Rhodotorula mucilaginosa dominated in the spruce forest and Cryptococcus magnus was more abundant in the swamp. Our study confirmed the low occurrence of tremellaceous yeasts in the Sphagnum phyllosphere. Of the few isolated ascomycetous yeast and yeast-like species, some were differentiated from hitherto known species in physiological tests and phylogenetic analyses. We describe one of them as Candida sphagnicola and designate KBP Y-3887(T) (=CBS 11774(T) = VKPM Y-3566(T) = MUCL 53590(T)) as the type strain. The new species was registered in MycoBank under MB 563443.

Non-Saccharomyces yeasts protect against epithelial cell barrier disruption induced by Salmonella enterica subsp. enterica serovar Typhimurium.

PubMed

Smith, I M; Baker, A; Arneborg, N; Jespersen, L

2015-11-01

The human gastrointestinal epithelium makes up the largest barrier separating the body from the external environment. Whereas invasive pathogens cause epithelial barrier disruption, probiotic micro-organisms modulate tight junction regulation and improve epithelial barrier function. In addition, probiotic strains may be able to reduce epithelial barrier disruption caused by pathogenic species. The aim of this study was to explore non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Benchmarking against established probiotic strains, we evaluated the ability of four nonpathogenic yeast species to modulate transepithelial electrical resistance (TER) across a monolayer of differentiated human colonocytes (Caco-2 cells). Further, we assessed yeast modulation of a Salmonella Typhimurium-induced epithelial cell barrier function insult. Our findings demonstrate distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function. While the established probiotic yeast Saccharomyces boulardii increased TER across a Caco-2 monolayer by 30%, Kluyveromyces marxianus exhibited significantly stronger properties of TER enhancement (50% TER increase). In addition, our data demonstrate significant yeast-mediated modulation of Salmonella-induced epithelial cell barrier disruption and identify K. marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. This study demonstrates distinct patterns of non-Saccharomyces yeast modulation of epithelial cell barrier function in vitro. Further, our data demonstrate significant yeast-mediated modulation of Salmonella Typhimurium-induced epithelial cell barrier disruption and identify Kluyveromyces marxianus and Metschnikowia gruessii as two non-Saccharomyces yeasts capable of protecting human epithelial cells from pathogen invasion. This study is the first to demonstrate significant non-Saccharomyces yeast-mediated epithelial cell barrier protection from Salmonella invasion, thus encouraging future efforts aimed at confirming the observed effects in vivo and driving further strain development towards novel yeast probiotics. © 2015 The Society for Applied Microbiology.

The yeast actin cytoskeleton.

PubMed

Mishra, Mithilesh; Huang, Junqi; Balasubramanian, Mohan K

2014-03-01

The actin cytoskeleton is a complex network of dynamic polymers, which plays an important role in various fundamental cellular processes, including maintenance of cell shape, polarity, cell division, cell migration, endocytosis, vesicular trafficking, and mechanosensation. Precise spatiotemporal assembly and disassembly of actin structures is regulated by the coordinated activity of about 100 highly conserved accessory proteins, which nucleate, elongate, cross-link, and sever actin filaments. Both in vivo studies in a wide range of organisms from yeast to metazoans and in vitro studies of purified proteins have helped shape the current understanding of actin dynamics and function. Molecular genetics, genome-wide functional analysis, sophisticated real-time imaging, and ultrastructural studies in concert with biochemical analysis have made yeast an attractive model to understand the actin cytoskeleton, its molecular dynamics, and physiological function. Studies of the yeast actin cytoskeleton have contributed substantially in defining the universal mechanism regulating actin assembly and disassembly in eukaryotes. Here, we review some of the important insights generated by the study of actin cytoskeleton in two important yeast models the budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Ferrocene-fused derivatives of acenes, tropones and thiepins

NASA Astrophysics Data System (ADS)

Maharjan, Bidhya Laxmi

This research project is concentrated on tuning the properties of small organic molecules, namely polyacenes, tropones and thiepins, by incorporating redox-active transition metal centers pi-bonded to terminal cyclopentadienyl ligands. Organometallicfused acenequinones, tropones, thiepins and cyclopentadiene-capped polyacenes were synthesized and characterized. This work was divided into three parts: first, the synthesis of ferrocene-fused acenequinones, cyclopentadiene-capped acenequinones and their subsequent aromatization to polyacenes; second, the synthesis of ferrocene-fused tropones, thiotropones and tropone oxime; and third, the synthesis of ferrocene-fused thiepins. Ferrocene-fused quinones are the precursors to our target complexes. Our synthetic route to ferrocenequinones involved two-fold aldol condensation between 1,2- diformylferrocene and naphthalene-1,4-diol or anthracene-1,4-diol, and four-fold condensation between 1,2-diformylferrocene and 1,4-cyclohexanedione. Reduction of ferrocene-fused quinones with borane in THF resulted in ferrocene-fused dihydroacenes. Attempts to reduce ferrocene-fused acenequinones with sodium dithionite led to metalfree cyclopentadiene- (Cp-) capped acenequinones. Cp-capped acenequinones were aromatized to bis(triisopropylsilyl)ethynyl polyacenes by using lithium (triisopropylsilyl)acetylide (TIPSC≡CLi) with subsequent dehydroxylation by stannous chloride. The compounds were characterized by using spectroscopic methods and X-ray crystallography. Further, the electronic properties of these compounds were studied by using cyclic voltammetry and UV-visible spectroscopy. Cyclic voltammetry showed oxidation potentials of Cp-capped TIPS-tetracene and bis-Cp-capped TIPS-anthracene as 0.49 V and 0.61 V, respectively (vs. ferrocene/ferrocenium). The electrochemical band gaps were 2.15 eV and 2.58 eV, respectively. Organic thin-film transistor device performance of Cp-capped polyacenes was studied using solution deposition bottomcontact, bottom-gate (BCBG) device architecture and the resulting performance parameters are described herein. Similarly, we are also interested in potential applications of metallocene-fused tropones and derivatives as organic electronic materials. Condensation of 1,2- diformylferrocene with acetone or 1,3-diphenylacetone in the presence of KOH resulted in the ferrocene-fused tropone (eta5-2,4-cyclopentadien-1-yl)[(1,2,3,3a,8a-eta)-1,6-dihydro- 6-oxo-1-azulenyl]iron (1, R = H, E = O) and its 5,7-diphenyl derivative (1, R = Ph, E = O) as previously reported by Tirouflet. The use of piperidine as base resulted in Michael addition of piperidine to one of the carbon-carbon double bonds of the tropones. Lawesson's reagent converted the ferrocene-fused tropones to either a thiotropone (1, R = H, E = S) or a detached 5,7-diphenylazulenethiol (2). Reaction of the ferrocene-fused thiotropone with hydroxylamine gave the corresponding oxime (1, R = H, E = NOH). Products were characterized by using spectroscopic methods and X-ray crystallography. Their electronic properties were studied by using cyclic voltammetry and UV-visible spectroscopy. The third project involved the two-fold aldol condensation of 1,2- diformylferrocene with dimethylthioglycolate S-oxide in the presence of freshly distilled triethylamine, which gave mono- and di-dehydrated products. Deoxygenation of the ferrocene-fused thiepin S-oxide with 2-chloro-1,3,2-benzodioxaphosphole in the presence of pyridine resulted in the corresponding thiepin. The ester groups of the thiepin and thiepin S-oxide were hydrolyzed under basic conditions to give carboxylic acids, which were converted into acid chlorides using oxalyl chloride. Attempts to decarboxylate the thiepin and thiepin S-oxide diacids resulted in decomposition.

Yeast as a tool to identify anti-aging compounds

PubMed Central

Zimmermann, Andreas; Hofer, Sebastian; Pendl, Tobias; Kainz, Katharina; Madeo, Frank; Carmona-Gutierrez, Didac

2018-01-01

Abstract In the search for interventions against aging and age-related diseases, biological screening platforms are indispensable tools to identify anti-aging compounds among large substance libraries. The budding yeast, Saccharomyces cerevisiae, has emerged as a powerful chemical and genetic screening platform, as it combines a rapid workflow with experimental amenability and the availability of a wide range of genetic mutant libraries. Given the amount of conserved genes and aging mechanisms between yeast and human, testing candidate anti-aging substances in yeast gene-deletion or overexpression collections, or de novo derived mutants, has proven highly successful in finding potential molecular targets. Yeast-based studies, for example, have led to the discovery of the polyphenol resveratrol and the natural polyamine spermidine as potential anti-aging agents. Here, we present strategies for pharmacological anti-aging screens in yeast, discuss common pitfalls and summarize studies that have used yeast for drug discovery and target identification. PMID:29905792

Nectar Yeasts in the Tall Larkspur Delphinium barbeyi (Ranunculaceae) and Effects on Components of Pollinator Foraging Behavior

PubMed Central

Schaeffer, Robert N.; Phillips, Cody R.; Duryea, M. Catherine; Andicoechea, Jonathan; Irwin, Rebecca E.

2014-01-01

Microorganisms frequently colonize the nectar of angiosperm species. Though capable of altering a suite of traits important for pollinator attraction, few studies exist that test the degree to which they mediate pollinator foraging behavior. The objective of our study was to fill this gap by assessing the abundance and diversity of yeasts associated with the perennial larkspur Delphinium barbeyi (Ranunculaceae) and testing whether their presence affected components of pollinator foraging behavior. Yeasts frequently colonized D. barbeyi nectar, populating 54–77% of flowers examined depending on site. Though common, the yeast community was species-poor, represented by a single species, Metschnikowia reukaufii. Female-phase flowers of D. barbeyi were more likely to have higher densities of yeasts in comparison to male-phase flowers. Pollinators were likely vectors of yeasts, as virgin (unvisited) flowers rarely contained yeasts compared to flowers open to pollinator visitation, which were frequently colonized. Finally, pollinators responded positively to the presence of yeasts. Bombus foragers both visited and probed more flowers inoculated with yeasts in comparison to uninoculated controls. Taken together, our results suggest that variation in the occurrence and density of nectar-inhabiting yeasts have the potential to alter components of pollinator foraging behavior linked to pollen transfer and plant fitness. PMID:25272164

Yeast: An Overlooked Component of Bactrocera tryoni (Diptera: Tephritidae) Larval Gut Microbiota.

PubMed

Deutscher, Ania T; Reynolds, Olivia L; Chapman, Toni A

2017-02-01

Yeasts, often in hydrolyzed form, are key ingredients in the larval and adult diets of tephritid fruit fly colonies. However, very little is known about the presence or role of yeasts in the diets of tephritid fruit flies in nature. Previous studies have identified bacteria but not detected yeasts in the gut of Queensland fruit fly, Bactrocera tryoni (Froggatt), one of Australia's most economically damaging insect pests of horticultural crops and of significant biosecurity concern domestically and internationally. Here we demonstrate that cultivable yeasts are commonly found in the gut of B. tryoni larvae from fruit hosts. Analysis of the ITS1, 5.8S rRNA gene, and ITS2 sequences of randomly selected isolates identified yeasts and yeast-like fungi of the genera Aureobasidium, Candida, Cryptococcus, Hanseniaspora, Pichia, and Starmerella. The prevalence of these yeasts in fruits suggests that larvae consume the yeasts as part of their diet. This work highlights that yeasts should be considered in future tephritid larval gut microbiota studies. Understanding tephritid-microbial symbiont interactions will lead to improvements in artificial diets and the quality of mass-reared tephritids for the sterile insect technique. © The Authors 2016. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Preparation of corncob grits as a carrier for immobilizing yeast cells for ethanol production.

PubMed

Lee, Sang-Eun; Lee, Choon Geun; Kang, Do Hyung; Lee, Hyeon-Yong; Jung, Kyung-Hwan

2012-12-01

In this study, DEAE-corncobs [delignified corncob grits derivatized with 2-(diethylamino)ethyl chloride hydrochloride (DEAE·HCl)] were prepared as a carrier to immobilize yeast (Saccharomyces cerevisiae) for ethanol production. The immobilized yeast cell reactor produced ethanol under optimized DEAE·HCl derivatization and adsorption conditions between yeast cells and the DEAE-corncobs. When delignified corncob grit (3.0 g) was derivatized with 0.5M DEAE·HCl, the yeast cell suspension (OD600 = 3.0) was adsorbed at >90% of the initial cell OD600. This amount of adsorbed yeast cells was estimated to be 5.36 mg-dry cells/g-DEAE corncobs. The Qmax (the maximum cell adsorption by the carrier) of the DEAE-corncobs was estimated to be 25.1 (mg/g), based on a Languir model biosorption isotherm experiment. When we conducted a batch culture with medium recycling using the immobilized yeast cells, the yeast cells on DEAE-corncobs produced ethanol gradually, according to glucose consumption, without cells detaching from the DEAE-corncobs. We observed under electron microscopy that the yeast cells grew on the surface and in the holes of the DEAEcorncobs. In a future study, DEAE-corncobs and the immobilized yeast cell reactor system will contribute to bioethanol production from biomass hydrolysates.

Self-healing fuse development

NASA Technical Reports Server (NTRS)

Jones, N. D.; Kinsinger, R. E.; Harris, L. P.

1973-01-01

The mercury-filled self-healing fuses developed for this program afford very good protection from circuit faults with rapid reclosure. Fuse performance and design parameters have been characterized. Life tests indicate a capability of 500 fuse operations. Fuse ratings are 150 v at 5, 15, 25 and 50 circuit A. A series of sample fuses using alumina and beryllia insulation have been furnished to NASA for circuit evaluation.

Characterization of the tribological coating composition 77 wt % CaF2 - 23 wt % Li F fused to IN-750 alloy

NASA Technical Reports Server (NTRS)

Deadmore, D. L.; Sliney, H. E.

1986-01-01

A coating composed of 77 wt % CaF2 - 23 wt % LiF fused on IN-750 nickel-based alloy was studied using SEM, XRD, EDX, and optical microscopic methods. The surfaces examined were the as-fused coating with no subsequent treatment, the coating after ultrasonic cleaning in water, and the uncoated polished and etched metal. It was found that the coating reacts during fusion with Ti and Nb rich inclusions in the alloy. Numerous small rectangular crystallites of Ca(Ti,Nb) oxide are formed beneath an overlay of fused fluoride composition. These crystallines are stubby and appear to be embedded in the metal substrate surface. It is known from previous studies that this coating-alloy system has good tribological properties in extreme conditions, such as liquid fluorine. It has been concluded from the present study that the short firmly embedded crystalline protuberances contribute to the coating adherence and thereby to enhanced coating wear life.

Experimental Systems to Study Yeast Pexophagy.

PubMed

Yamashita, Shun-Ichi; Oku, Masahide; Sakai, Yasuyoshi; Fujiki, Yukio

2017-01-01

Peroxisome abundance is tightly regulated according to the physiological contexts, through regulations of both proliferation and degradation of the organelles. Here, we describe detailed methods to analyze processes for autophagic degradation of peroxisomes, termed pexophagy, in yeast organisms. The assay systems include a method for biochemical detection of pexophagy completion, and one for microscopic visualization of specialized membrane structures acting in pexophagy. As a model yeast organism utilized in studies of pexophagy, the methylotrophic yeast Komagataella phaffii (Pichia pastoris) is referred to in this chapter and related information on the studies with baker's yeast (Saccharomyces cerevisiae) is also included. The described techniques facilitate elucidation of molecular machineries for pexophagy and understanding of peroxisome-selective autophagic pathways.

Functional adaptation between yeast actin and its cognate myosin motors.

PubMed

Stark, Benjamin C; Wen, Kuo-Kuang; Allingham, John S; Rubenstein, Peter A; Lord, Matthew

2011-09-02

We employed budding yeast and skeletal muscle actin to examine the contribution of the actin isoform to myosin motor function. While yeast and muscle actin are highly homologous, they exhibit different charge density at their N termini (a proposed myosin-binding interface). Muscle myosin-II actin-activated ATPase activity is significantly higher with muscle versus yeast actin. Whether this reflects inefficiency in the ability of yeast actin to activate myosin is not known. Here we optimized the isolation of two yeast myosins to assess actin function in a homogenous system. Yeast myosin-II (Myo1p) and myosin-V (Myo2p) accommodate the reduced N-terminal charge density of yeast actin, showing greater activity with yeast over muscle actin. Increasing the number of negative charges at the N terminus of yeast actin from two to four (as in muscle) had little effect on yeast myosin activity, while other substitutions of charged residues at the myosin interface of yeast actin reduced activity. Thus, yeast actin functions most effectively with its native myosins, which in part relies on associations mediated by its outer domain. Compared with yeast myosin-II and myosin-V, muscle myosin-II activity was very sensitive to salt. Collectively, our findings suggest differing degrees of reliance on electrostatic interactions during weak actomyosin binding in yeast versus muscle. Our study also highlights the importance of native actin isoforms when considering the function of myosins.

Biochemical and structural characterization of a novel family of cystathionine beta-synthase domain proteins fused to a Zn ribbon-like domain.

PubMed

Proudfoot, Michael; Sanders, Stephen A; Singer, Alex; Zhang, Rongguang; Brown, Greg; Binkowski, Andrew; Xu, Linda; Lukin, Jonathan A; Murzin, Alexey G; Joachimiak, Andrzej; Arrowsmith, Cheryl H; Edwards, Aled M; Savchenko, Alexei V; Yakunin, Alexander F

2008-01-04

We have identified a novel family of proteins, in which the N-terminal cystathionine beta-synthase (CBS) domain is fused to the C-terminal Zn ribbon domain. Four proteins were overexpressed in Escherichia coli and purified: TA0289 from Thermoplasma acidophilum, TV1335 from Thermoplasma volcanium, PF1953 from Pyrococcus furiosus, and PH0267 from Pyrococcus horikoshii. The purified proteins had a red/purple color in solution and an absorption spectrum typical of rubredoxins (Rds). Metal analysis of purified proteins revealed the presence of several metals, with iron and zinc being the most abundant metals (2-67% of iron and 12-74% of zinc). Crystal structures of both mercury- and iron-bound TA0289 (1.5-2.0 A resolution) revealed a dimeric protein whose intersubunit contacts are formed exclusively by the alpha-helices of two cystathionine beta-synthase subdomains, whereas the C-terminal domain has a classical Zn ribbon planar architecture. All proteins were reversibly reduced by chemical reductants (ascorbate or dithionite) or by the general Rd reductase NorW from E. coli in the presence of NADH. Reduced TA0289 was found to be capable of transferring electrons to cytochrome C from horse heart. Likewise, the purified Zn ribbon protein KTI11 from Saccharomyces cerevisiae had a purple color in solution and an Rd-like absorption spectrum, contained both iron and zinc, and was reduced by the Rd reductase NorW from E. coli. Thus, recombinant Zn ribbon domains from archaea and yeast demonstrate an Rd-like electron carrier activity in vitro. We suggest that, in vivo, some Zn ribbon domains might also bind iron and therefore possess an electron carrier activity, adding another physiological role to this large family of important proteins.

Evidence that the metabolite repair enzyme NAD(P)HX epimerase has a moonlighting function.

PubMed

Niehaus, Thomas D; Elbadawi-Sidhu, Mona; Huang, Lili; Prunetti, Laurence; Gregory, Jesse F; de Crécy-Lagard, Valérie; Fiehn, Oliver; Hanson, Andrew D

2018-06-29

NAD(P)H-hydrate epimerase (EC 5.1.99.6) is known to help repair NAD(P)H hydrates (NAD(P)HX), which are damage products existing as R and S epimers. The S epimer is reconverted to NAD(P)H by a dehydratase; the epimerase facilitates epimer interconversion. Epimerase deficiency in humans causes a lethal disorder attributed to NADHX accumulation. However, bioinformatic evidence suggest caution about this attribution by predicting that the epimerase has a second function connected to vitamin B 6 (pyridoxal 5'-phosphate and related compounds). Specifically, (i) the epimerase is fused to a B 6 salvage enzyme in plants, (ii) epimerase genes cluster on the chromosome with B 6 -related genes in bacteria, and (iii) epimerase and B 6 -related genes are coexpressed in yeast and Arabidopsis The predicted second function was explored in Escherichia coli , whose epimerase and dehydratase are fused and encoded by yjeF The putative NAD(P)HX epimerase active site has a conserved lysine residue (K192 in E. coli YjeF). Changing this residue to alanine cut in vitro epimerase activity by ≥95% but did not affect dehydratase activity. Mutant cells carrying the K192A mutation had essentially normal NAD(P)HX dehydratase activity and NAD(P)HX levels, showing that the mutation had little impact on NAD(P)HX repair in vivo However, these cells showed metabolome changes, particularly in amino acids, which exceeded those in cells lacking the entire yjeF gene. The K192A mutant cells also had reduced levels of 'free' (i.e. weakly bound or unbound) pyridoxal 5'-phosphate. These results provide circumstantial evidence that the epimerase has a metabolic function beyond NAD(P)HX repair and that this function involves vitamin B 6 . © 2018 The Author(s).

Evidence that the metabolite repair enzyme NAD(P)HX epimerase has a moonlighting function

PubMed Central

Niehaus, Thomas D.; Elbadawi-Sidhu, Mona; Huang, Lili; Prunetti, Laurence; Gregory, Jesse F.; de Crécy-Lagard, Valérie; Fiehn, Oliver; Hanson, Andrew D.

2018-01-01

NAD(P)H-hydrate epimerase (EC 5.1.99.6) is known to help repair NAD(P)H hydrates (NAD(P)HX), which are damage products existing as R and S epimers. The S epimer is reconverted to NAD(P)H by a dehydratase; the epimerase facilitates epimer interconversion. Epimerase deficiency in humans causes a lethal disorder attributed to NADHX accumulation. However, bioinformatic evidence suggest caution about this attribution by predicting that the epimerase has a second function connected to vitamin B6 (pyridoxal 5′-phosphate and related compounds). Specifically, (i) the epimerase is fused to a B6 salvage enzyme in plants, (ii) epimerase genes cluster on the chromosome with B6-related genes in bacteria, and (iii) epimerase and B6-related genes are coexpressed in yeast and Arabidopsis. The predicted second function was explored in Escherichia coli, whose epimerase and dehydratase are fused and encoded by yjeF. The putative NAD(P)HX epimerase active site has a conserved lysine residue (K192 in E. coli YjeF). Changing this residue to alanine cut in vitro epimerase activity by ≥95% but did not affect dehydratase activity. Mutant cells carrying the K192A mutation had essentially normal NAD(P)HX dehydratase activity and NAD(P)HX levels, showing that the mutation had little impact on NAD(P)HX repair in vivo. However, these cells showed metabolome changes, particularly in amino acids, which exceeded those in cells lacking the entire yjeF gene. The K192A mutant cells also had reduced levels of ‘free’ (i.e. weakly bound or unbound) pyridoxal 5'-phosphate. These results provide circumstantial evidence that the epimerase has a metabolic function beyond NAD(P)HX repair and that this function involves vitamin B6. PMID:29654173

Interactions of Condensed Tannins with Saccharomyces cerevisiae Yeast Cells and Cell Walls: Tannin Location by Microscopy.

PubMed

Mekoue Nguela, Julie; Vernhet, Aude; Sieczkowski, Nathalie; Brillouet, Jean-Marc

2015-09-02

Interactions between grape tannins/red wine polyphenols and yeast cells/cell walls was previously studied within the framework of red wine aging and the use of yeast-derived products as an alternative to aging on lees. Results evidenced a quite different behavior between whole cells (biomass grown to elaborate yeast-derived products, inactivated yeast, and yeast inactivated after autolysis) and yeast cell walls (obtained from mechanical disruption of the biomass). Briefly, whole cells exhibited a high capacity to irreversibly adsorb grape and wine tannins, whereas only weak interactions were observed for cell walls. This last point was quite unexpected considering the literature and called into question the real role of cell walls in yeasts' ability to fix tannins. In the present work, tannin location after interactions between grape and wine tannins and yeast cells and cell walls was studied by means of transmission electron microscopy, light epifluorescence, and confocal microscopy. Microscopy observations evidenced that if tannins interact with cell walls, and especially cell wall mannoproteins, they also diffuse freely through the walls of dead cells to interact with their plasma membrane and cytoplasmic components.

Fused Bead Analysis of Diogenite Meteorites

NASA Technical Reports Server (NTRS)

Mittlefehldt, D.W.; Beck, B.W.; McSween, H.Y.; Lee, C.T. A.

2009-01-01

Bulk rock chemistry is an essential dataset in meteoritics and planetary science [1]. A common method used to obtain the bulk chemistry of meteorites is ICP-MS. While the accuracy, precision and low detection limits of this process are advantageous [2], the sample size used for analysis (approx.70 mg) can be a problem in a field where small and finite samples are the norm. Fused bead analysis is another bulk rock analytical technique that has been used in meteoritics [3]. This technique involves forming a glass bead from 10 mg of sample and measuring its chemistry using a defocused beam on a microprobe. Though the ICP-MS has lower detection limits than the microprobe, the fused bead method destroys a much smaller sample of the meteorite. Fused bead analysis was initially designed for samples with near-eutectic compositions and low viscosities. Melts generated of this type homogenize at relatively low temperatures and produce primary melts near the sample s bulk composition [3]. The application of fused bead analysis to samples with noneutectic melt compositions has not been validated. The purpose of this study is to test if fused bead analysis can accurately determine the bulk rock chemistry of non-eutectic melt composition meteorites. To determine this, we conduct two examinations of the fused bead. First, we compare ICP-MS and fused bead results of the same samples using statistical analysis. Secondly, we inspect the beads for the presence of crystals and chemical heterogeneity. The presence of either of these would indicate incomplete melting and quenching of the bead.

Yeast diversity associated to sediments and water from two Colombian artificial lakes

PubMed Central

Silva-Bedoya, L.M.; Ramírez-Castrillón, M.; Osorio-Cadavid, E.

2014-01-01

In Colombia, knowledge of the yeast and yeast-like fungi community is limited because most studies have focused on species with clinical importance. Sediments and water represent important habitats for the study of yeast diversity, especially for yeast species with industrial, biotechnological, and bioremediation potential. The main purpose of this study was to identify and compare the diversity of yeast species associated with sediment and water samples from two artificial lakes in Universidad del Valle (Cali-Colombia). Yeast samplings were performed from fifteen sediment samples and ten water samples. Grouping of similar isolates was initially based on colony and cell morphology, which was then complemented by micro/mini satellite primed PCR banding pattern analysis by using GTG5 as single primer. A representative isolate for each group established was chosen for D1/D2 domain sequencing and identification. In general, the following yeast species were identified: Candida albicans, Candida diversa, Candida glabrata, Candida pseudolambica, Cryptococcus podzolicus, Cryptococcus rajasthanensis, Cryptococcus laurentii, Williopsis saturnus, Hanseniaspora thailandica, Hanseniaspora uvarum, Rhodotorula mucilaginosa, Saccharomyces cerevisiae, Torulaspora delbrueckii, Torulaspora pretoriensis, Tricosporon jirovecii, Trichosporon laibachii and Yarrowia lypolitica. Two possible new species were also found, belonging to the Issatchenkia sp. and Bullera sp. genera. In conclusion, the lakes at the Universidad del Valle campus have significant differences in yeast diversity and species composition between them. PMID:24948924

Yeast species associated with wine grapes in China.

PubMed

Li, Shuang-Shi; Cheng, Chao; Li, Zheng; Chen, Jing-Yu; Yan, Bin; Han, Bei-Zhong; Reeves, Malcolm

2010-03-31

Having more information on the yeast ecology of grapes is important for wine-makers to produce wine with high quality and typical attributes. China is a significant wine-consuming country and is becoming a serious wine-producer, but little has been reported about the yeast ecology of local ecosystems. This study provides the first step towards the exploitation of the yeast wealth in China's vine-growing regions. The aim of this study was to investigate the yeast population density and diversity on three grape varieties cultivated in four representative vine-growing regions of China. Yeast species diversity was evaluated by using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) and sequence analysis of the 5.8S internal transcribed spacer (ITS) ribosomal DNA (rDNA) region of cultivable yeasts. The grapes harbored yeast populations at 10(2)-10(6)CFU/mL, consisting mostly of non-Saccharomyces species. Seventeen different yeast species belonging to eight genera were detected on the grape samples tested, including Hanseniaspora uvarum, Cryptococcus flavescens, Pichia fermentans, Candida zemplinina, Cryptococcus carnescens, Candida inconpicua, Zygosaccharomyces fermentati, Issatchenkia terricola, Candida quercitrusa, Hanseniaspora guilliermondii, Candida bombi, Zygosaccharomyces bailii, Sporidiobolus pararoseus, Cryptococcus magnus, Metschnikowia pulcherrima, Issatchenkia orientalis and Pichia guilliermondii. H. uvarum and C. flavescens were the dominant species present on the grapes. For the first time Sporidiobolus pararoseus was discovered as an inhabitant of the grape ecosystem. The yeast community on grape berries was influenced by the grape chemical composition, vine-variety and vine-growing region. This study is the first to identify the yeast communities associated with grapes in China using molecular methods. The results enrich our knowledge of wine-related microorganisms, and can be used to promote the development of the local wine industry. Copyright 2010 Elsevier B.V. All rights reserved.

Ultraviolet Laser Damage Dependence on Contamination Concentration in Fused Silica Optics during Reactive Ion Etching Process

PubMed Central

Sun, Laixi; Shao, Ting; Shi, Zhaohua; Huang, Jin; Ye, Xin; Jiang, Xiaodong; Wu, Weidong; Yang, Liming; Zheng, Wanguo

2018-01-01

The reactive ion etching (RIE) process of fused silica is often accompanied by surface contamination, which seriously degrades the ultraviolet laser damage performance of the optics. In this study, we find that the contamination behavior on the fused silica surface is very sensitive to the RIE process which can be significantly optimized by changing the plasma generating conditions such as discharge mode, etchant gas and electrode material. Additionally, an optimized RIE process is proposed to thoroughly remove polishing-introduced contamination and efficiently prevent the introduction of other contamination during the etching process. The research demonstrates the feasibility of improving the damage performance of fused silica optics by using the RIE technique. PMID:29642571

Materials Testing and Cost Modeling for Composite Parts Through Additive Manufacturing

DTIC Science & Technology

2016-04-30

FDM include plastic jet printing (PJP), fused filament modeling ( FFM ), and fused filament fabrication (FFF). FFF was coined by the RepRap project to...additive manufacturing processes? • Fused deposition modeling (FDM) trademarked by Stratasys • Fused filament modeling ( FFM ) and fused filament

Observation of changes in the metallurgical characteristics of Ni-Cr alloys using Nd:YAG laser welding

PubMed Central

Choi, SM

2014-01-01

This study aimed to determine the effect of hardness change according to penetration depth in the laser fusing zone and observed the correlation of the microstructure as an Nd:YAG laser was irradiated to Ni-Cr alloy for dental use by setting the spot diameter size to various conditions. In all groups, the hardness depth profiles in the laser fusing zone and heat-affected zone (HAZ) had larger values than those of the base metal. In addition, the hardness values in places beyond the fusing zone and the HAZ were measured as being quantitatively lower. The observation result of the diffusion of the constituent elements and microstructure using field emission scanning electron microscopy, energy-dispersive spectroscopy, and electron probe microanalyzer showed that the fusing zone revealed a much finer dendritic form than the base metal due to the self-quenching effect after welding, while no change in constituent elements was found although some evaporation of the main elements was observed. In addition, Mo- and Si-combined intermetallic compounds were formed on the interdendritic area. Through this study, the laser fusing zone had better hardenability due to the intermetallic compound and grain refinement effect. PMID:25342985

The presence of Enterococcus, coliforms and E. coli in a commercial yeast manufacturing process.

PubMed

O'Brien, S S; Lindsay, D; von Holy, A

2004-07-01

This study evaluated a typical commercial yeast manufacturing process for bacterial contamination. Product line samples of a commercial yeast manufacturing process and the corresponding seed yeast manufacturing process were obtained upstream from the final compressed and dry yeast products. All samples were analysed before (non-PI) and after preliminary incubation (PI) at 37 degrees C for 24 h. The PI procedure was incorporated for amplification of bacterial counts below the lower detection limit. Enterococcus, coliform and Escherichia coli counts were quantified by standard pour-plate techniques using selective media. Presence at all stages and progressive increases in counts of Enterococcus, coliforms and E. coli during processing in the commercial manufacturing operation suggested that the primary source of contamination of both compressed and dry yeast with these bacteria was the seed yeast manufacturing process and that contamination was amplified throughout the commercial yeast manufacturing process. This was confirmed by surveys of the seed yeast manufacturing process which indicated that contamination of the seed yeast with Enterococcus, coliforms and E. coli occurred during scale up of seed yeast biomass destined as inoculum for the commercial fermentation.

Screening of the five different wild, traditional and industrial Saccharomyces cerevisiae strains to overproduce bioethanol in the batch submerged fermentation.

PubMed

Shaghaghi-Moghaddam, Reza; Jafarizadeh-Malmiri, Hoda; Mehdikhani, Parviz; Jalalian, Sepide; Alijanianzadeh, Reza

2017-12-28

Efforts to produce bioethanol with higher productivity in a batch submerged fermentation were made by evaluating the bioethanol production of the five different strains of Saccharomyces cerevisiae, namely, NCYC 4109 (traditional bakery yeast), SFO6 (industrial yeast), TTCC 2956 (hybrid baking yeast) and two wild yeasts, PTCC 5052 and BY 4743. The bioethanol productivity and kinetic parameters for all five yeasts at constant fermentation conditions, during 72 h, were evaluated and monitored. The obtained results indicated that compared to the wild yeasts, both traditional bakery (NCYC 4109) and industrial (SFO6) yeasts had higher bioethanol productivity (0.9 g/L h). Significant (p<0.05) differences between biomass concentration of NCYC 4109 yeast and those of other yeasts 30 h after start of fermentation, and its high bioethanol concentration (59.19 g/L) and yield over consumed sugars (77.25%) were highlighted among all the studied yeasts. Minimum bioethanol productivity was obtained using yeasts PTCC 5052 (0.7 g/L h) and TTCC 2956 (0.86 g/L h). However, maximum yield over consumed sugar was obtained using the yeast TTCC 2956 (79.41%).

Production, characterization and gene cloning of the extracellular enzymes from the marine-derived yeasts and their potential applications.

PubMed

Chi, Zhenming; Chi, Zhe; Zhang, Tong; Liu, Guanglei; Li, Jing; Wang, Xianghong

2009-01-01

In this review article, the extracellular enzymes production, their properties and cloning of the genes encoding the enzymes from marine yeasts are overviewed. Several yeast strains which could produce different kinds of extracellular enzymes were selected from the culture collection of marine yeasts available in this laboratory. The strains selected belong to different genera such as Yarrowia, Aureobasidium, Pichia, Metschnikowia and Cryptococcus. The extracellular enzymes include cellulase, alkaline protease, aspartic protease, amylase, inulinase, lipase and phytase, as well as killer toxin. The conditions and media for the enzyme production by the marine yeasts have been optimized and the enzymes have been purified and characterized. Some genes encoding the extracellular enzymes from the marine yeast strains have been cloned, sequenced and expressed. It was found that some properties of the enzymes from the marine yeasts are unique compared to those of the homologous enzymes from terrestrial yeasts and the genes encoding the enzymes in marine yeasts are different from those in terrestrial yeasts. Therefore, it is of very importance to further study the enzymes and their genes from the marine yeasts. This is the first review on the extracellular enzymes and their genes from the marine yeasts.

An Ancient Yeast for Young Geneticists: A Primer on the Schizosaccharomyces pombe Model System.

PubMed

Hoffman, Charles S; Wood, Valerie; Fantes, Peter A

2015-10-01

The fission yeast Schizosaccharomyces pombe is an important model organism for the study of eukaryotic molecular and cellular biology. Studies of S. pombe, together with studies of its distant cousin, Saccharomyces cerevisiae, have led to the discovery of genes involved in fundamental mechanisms of transcription, translation, DNA replication, cell cycle control, and signal transduction, to name but a few processes. However, since the divergence of the two species approximately 350 million years ago, S. pombe appears to have evolved less rapidly than S. cerevisiae so that it retains more characteristics of the common ancient yeast ancestor, causing it to share more features with metazoan cells. This Primer introduces S. pombe by describing the yeast itself, providing a brief description of the origins of fission yeast research, and illustrating some genetic and bioinformatics tools used to study protein function in fission yeast. In addition, a section on some key differences between S. pombe and S. cerevisiae is included for readers with some familiarity with budding yeast research but who may have an interest in developing research projects using S. pombe. Copyright © 2015 by the Genetics Society of America.

STS-115 MS MacLean holds Yeast GAP in the FWD MDDK of the Space Shuttle Atlantis during Joint Operations

NASA Image and Video Library

2006-09-19

S115-E-07273 (9-21 Sept. 2006) --- Astronaut Heidemarie M. Stefanyshyn-Piper, STS-115 mission specialist, works with the Yeast-Group Activation Packs (Yeast-GAP) on the middeck of Space Shuttle Atlantis. Yeast-GAP experiment studies the effects of genetic changes of yeast cells exposed to the space environment. The results will help scientists to understand how cells respond to radiation and microgravity.

STS-115 MS MacLean holds Yeast GAP in the FWD MDDK of the Space Shuttle Atlantis during Joint Operations

NASA Image and Video Library

2006-09-19

S115-E-07274 (9-21 Sept. 2006) --- Astronaut Heidemarie M. Stefanyshyn-Piper, STS-115 mission specialist, works with the Yeast-Group Activation Packs (Yeast-GAP) on the middeck of Space Shuttle Atlantis. Yeast-GAP experiment studies the effects of genetic changes of yeast cells exposed to the space environment. The results will help scientists to understand how cells respond to radiation and microgravity.

Biotechnology of non-Saccharomyces yeasts-the basidiomycetes.

PubMed

Johnson, Eric A

2013-09-01

Yeasts are the major producer of biotechnology products worldwide, exceeding production in capacity and economic revenues of other groups of industrial microorganisms. Yeasts have wide-ranging fundamental and industrial importance in scientific, food, medical, and agricultural disciplines (Fig. 1). Saccharomyces is the most important genus of yeast from fundamental and applied perspectives and has been expansively studied. Non-Saccharomyces yeasts (non-conventional yeasts) including members of the Ascomycetes and Basidiomycetes also have substantial current utility and potential applicability in biotechnology. In an earlier mini-review, "Biotechnology of non-Saccharomyces yeasts-the ascomycetes" (Johnson Appl Microb Biotechnol 97: 503-517, 2013), the extensive biotechnological utility and potential of ascomycetous yeasts are described. Ascomycetous yeasts are particularly important in food and ethanol formation, production of single-cell protein, feeds and fodder, heterologous production of proteins and enzymes, and as model and fundamental organisms for the delineation of genes and their function in mammalian and human metabolism and disease processes. In contrast, the roles of basidiomycetous yeasts in biotechnology have mainly been evaluated only in the past few decades and compared to the ascomycetous yeasts and currently have limited industrial utility. From a biotechnology perspective, the basidiomycetous yeasts are known mainly for the production of enzymes used in pharmaceutical and chemical synthesis, for production of certain classes of primary and secondary metabolites such as terpenoids and carotenoids, for aerobic catabolism of complex carbon sources, and for bioremediation of environmental pollutants and xenotoxicants. Notwithstanding, the basidiomycetous yeasts appear to have considerable potential in biotechnology owing to their catabolic utilities, formation of enzymes acting on recalcitrant substrates, and through the production of unique primary and secondary metabolites. This and the earlier mini-review (Johnson Appl Microb Biotechnol 97:503-517, 2013) were motivated during the preparation and publication of the landmark three-volume set of "The yeasts: a taxonomic study, 5th edition" (Kurtzman et al. 2011a, b).

Persistence of Two Non-Saccharomyces Yeasts (Hanseniaspora and Starmerella) in the Cellar

PubMed Central

Grangeteau, Cédric; Gerhards, Daniel; von Wallbrunn, Christian; Alexandre, Hervé; Rousseaux, Sandrine; Guilloux-Benatier, Michèle

2016-01-01

Different genera and/or species of yeasts present on grape berries, in musts and wines are widely described. Nevertheless, the community of non-Saccharomyces yeasts present in the cellar is still given little attention. Thus it is not known if the cellar is a real ecological niche for these yeasts or if it is merely a transient habitat for populations brought in by grape berries during the winemaking period. This study focused on three species of non-Saccharomyces yeasts commonly encountered during vinification: Starmerella bacillaris (synonymy with Candida zemplinina), Hanseniaspora guilliermondii and Hanseniaspora uvarum. More than 1200 isolates were identified at the strain level by FT-IR spectroscopy (207 different FTIR strain pattern). Only a small proportion of non-Saccharomyces yeasts present in musts came directly from grape berries for the three species studied. Some strains were found in the must in two consecutive years and some of them were also found in the cellar environment before the arrival of the harvest of second vintage. This study demonstrates for the first time the persistence of non-Saccharomyces yeast strains from year to year in the cellar. Sulfur dioxide can affect yeast populations in the must and therefore their persistence in the cellar environment. PMID:27014199

Experimental Study of Membrane Fouling during Crossflow Microfiltration of Yeast and Bacteria Suspensions: Towards an Analysis at the Microscopic Level

PubMed Central

Ben Hassan, Ines; Ennouri, Monia; Lafforgue, Christine; Schmitz, Philippe; Ayadi, Abdelmoneim

2013-01-01

Microfiltration of model cell suspensions combining macroscopic and microscopic approaches was studied in order to better understand microbial membrane fouling mechanisms. The respective impact of Saccharomyces cerevisiae yeast and Escherichia coli bacteria on crossflow microfiltration performances was investigated using a multichannel ceramic 0.2 µm membrane. Pure yeast suspensions (5 µm ovoid cells) and mixtures of yeast and bacteria (1 to 2.5 µm rod shape cells) were considered in order to analyse the effect of interaction between these two microorganisms on fouling reversibility. The resistances varied significantly with the concentration and characteristics of the microorganisms. Membrane fouling with pure yeast suspension was mainly reversible. For yeast and bacteria mixed suspensions (6 g L−1 yeast concentration) the increase in bacteria from 0.15 to 0.30 g L−1 increased the percentage of normalized reversible resistance. At 10 g L−1 yeast concentration, the addition of bacteria tends to increase the percentage of normalized irreversible resistance. For the objective of performing local analysis of fouling, an original filtration chamber allowing direct in situ observation of the cake by confocal laser scanning microscopy (CLSM) was designed, developed and validated. This device will be used in future studies to characterize cake structure at the microscopic scale. PMID:24958619

Fusion to Human Serum Albumin Extends the Circulatory Half-Life and Duration of Antithrombotic Action of the Kunitz Protease Inhibitor Domain of Protease Nexin 2.

PubMed

Sheffield, William P; Eltringham-Smith, Louise J; Bhakta, Varsha

2018-01-01

The Kunitz Protease Inhibitor (KPI) domain of protease nexin 2 (PN2) potently inhibits coagulation factor XIa. Recombinant KPI has been shown to inhibit thrombosis in mouse models, but its clearance from the murine circulation remains uncharacterized. The present study explored the pharmacokinetic and pharmacodynamic effects of fusing KPI to human serum albumin (HSA) in fusion protein KPIHSA. Hexahistidine-tagged KPI (63 amino acids) and KPIHSA (656 amino acids) were expressed in Pichia pastoris yeast and purified by nickel-chelate chromatography. Clearance profiles in mice were determined, as well as the effects of KPI or KPIHSA administration on FeCl3-induced vena cava thrombus size or carotid artery time to occlusion, respectively. Fusion to HSA increased the mean terminal half-life of KPI by 8-fold and eliminated its interaction with the low density lipoprotein receptor-related protein. KPI and KPIHSA similarly reduced thrombus size and occlusion in both venous and arterial thrombosis models when administered at the time of injury, but only KPI was effective when administered one hour before injury. Albumin fusion deflects KPI from rapid in vivo clearance without impairing its antithrombotic properties and widens its potential therapeutic window. © 2018 The Author(s). Published by S. Karger AG, Basel.

Dissecting Stop Transfer versus Conservative Sorting Pathways for Mitochondrial Inner Membrane Proteins in Vivo*

PubMed Central

Park, Kwangjin; Botelho, Salomé Calado; Hong, Joonki; Österberg, Marie; Kim, Hyun

2013-01-01

Mitochondrial inner membrane proteins that carry an N-terminal presequence are sorted by one of two pathways: stop transfer or conservative sorting. However, the sorting pathway is known for only a small number of proteins, in part due to the lack of robust experimental tools with which to study. Here we present an approach that facilitates determination of inner membrane protein sorting pathways in vivo by fusing a mitochondrial inner membrane protein to the C-terminal part of Mgm1p containing the rhomboid cleavage region. We validated the Mgm1 fusion approach using a set of proteins for which the sorting pathway is known, and determined sorting pathways of inner membrane proteins for which the sorting mode was previously uncharacterized. For Sdh4p, a multispanning membrane protein, our results suggest that both conservative sorting and stop transfer mechanisms are required for insertion. Furthermore, the sorting process of Mgm1 fusion proteins was analyzed under different growth conditions and yeast mutant strains that were defective in the import motor or the m-AAA protease function. Our results show that the sorting of mitochondrial proteins carrying moderately hydrophobic transmembrane segments is sensitive to cellular conditions, implying that mitochondrial import and membrane sorting in the physiological environment may be dynamically tuned. PMID:23184936

Characterization of the Drosophila Methoprene -tolerant gene product. Juvenile hormone binding and ligand-dependent gene regulation.

PubMed

Miura, Ken; Oda, Masahito; Makita, Sumiko; Chinzei, Yasuo

2005-03-01

Juvenile hormones (JHs) of insects are sesquiterpenoids that regulate a great diversity of processes in development and reproduction. As yet the molecular modes of action of JH are poorly understood. The Methoprene-tolerant (Met) gene of Drosophila melanogaster has been found to be responsible for resistance to a JH analogue (JHA) insecticide, methoprene. Previous studies on Met have implicated its involvement in JH signaling, although direct evidence is lacking. We have now examined the product of Met (MET) in terms of its binding to JH and ligand-dependent gene regulation. In vitro synthesized MET directly bound to JH III with high affinity (Kd = 5.3 +/- 1.5 nm, mean +/- SD), consistent with the physiological JH concentration. In transient transfection assays using Drosophila S2 cells the yeast GAL4-DNA binding domain fused to MET exerted JH- or JHA-dependent activation of a reporter gene. Activation of the reporter gene was highly JH- or JHA-specific with the order of effectiveness: JH III > JH II > JH I > methoprene; compounds which are only structurally related to JH or JHA did not induce any activation. Localization of MET in the S2 cells was nuclear irrespective of the presence or absence of JH. These results suggest that MET may function as a JH-dependent transcription factor.

Making connections for life: an in vivo map of the yeast interactome.

PubMed

Kast, Juergen

2008-10-01

Proteins are the true workhorses of any cell. To carry out specific tasks, they frequently bind other molecules in their surroundings. Due to their structural complexity and flexibility, the most diverse array of interactions is seen with other proteins. The different geometries and affinities available for such interactions typically bestow specific functions on proteins. Having available a map of protein-protein interactions is therefore of enormous importance for any researcher interested in gaining insight into biological systems at the level of cells and organisms. In a recent report, a novel approach has been employed that relies on the spontaneous folding of complementary enzyme fragments fused to two different proteins to test whether these interact in their actual cellular context [Tarassov et al., Science 320, 1465-1470 (2008)]. Genome-wide application of this protein-fragment complementation assay has resulted in the first map of the in vivo interactome of Saccharomyces cerevisiae. The current data show striking similarities but also significant differences to those obtained using other large-scale approaches for the same task. This warrants a general discussion of the current state of affairs of protein-protein interaction studies and foreseeable future trends, highlighting their significance for a variety of applications and their potential to revolutionize our understanding of the architecture and dynamics of biological systems.

Making connections for life: an in vivo map of the yeast interactome

PubMed Central

Kast, Juergen

2008-01-01

Proteins are the true workhorses of any cell. To carry out specific tasks, they frequently bind other molecules in their surroundings. Due to their structural complexity and flexibility, the most diverse array of interactions is seen with other proteins. The different geometries and affinities available for such interactions typically bestow specific functions on proteins. Having available a map of protein–protein interactions is therefore of enormous importance for any researcher interested in gaining insight into biological systems at the level of cells and organisms. In a recent report, a novel approach has been employed that relies on the spontaneous folding of complementary enzyme fragments fused to two different proteins to test whether these interact in their actual cellular context [Tarassov et al., Science 320, 1465–1470 (2008)]. Genome-wide application of this protein-fragment complementation assay has resulted in the first map of the in vivo interactome of Saccharomyces cerevisiae. The current data show striking similarities but also significant differences to those obtained using other large-scale approaches for the same task. This warrants a general discussion of the current state of affairs of protein–protein interaction studies and foreseeable future trends, highlighting their significance for a variety of applications and their potential to revolutionize our understanding of the architecture and dynamics of biological systems. PMID:19404434

The membrane-tethered transcription factor ANAC089 serves as redox-dependent suppressor of stromal ascorbate peroxidase gene expression

PubMed Central

Klein, Peter; Seidel, Thorsten; Stöcker, Benedikt; Dietz, Karl-Josef

2012-01-01

The stromal ascorbate peroxidase (sAPX) functions as central element of the chloroplast antioxidant defense system. Its expression is under retrograde control of chloroplast signals including redox- and reactive oxygen species-linked cues. The sAPX promoter of Arabidopsis thaliana was dissected in transient reporter assays using mesophyll protoplasts. The study revealed regulatory elements up to –1868 upstream of the start codon. By yeast-one-hybrid screening, the transcription factor ANAC089 was identified to bind to the promoter fragment 2 (–1262 to –1646 bp upstream of translational initiation). Upon mutation of the cis-acting element CACG, binding of ANAC089 was abolished. Expression of a fused fluorescent protein version and comparison with known endomembrane markers localized ANAC089 to the trans-Golgi network and the ER. The transcription factor was released upon treatment with reducing agents and targeted to the nucleus. Transactivation assays using wild type and mutated versions of the promoter showed a partial suppression of reporter expression. The data indicate that ANAC089 functions in a negative retrograde loop, lowering sAPX expression if the cell encounters a highly reducing condition. This conclusion was supported by reciprocal transcript accumulation of ANAC089 and sAPX during acclimation to low, normal, and high light conditions. PMID:23162559

A Comparison of the Beneficial Effects of Live and Heat-Inactivated Baker's Yeast on Nile Tilapia: Suggestions on the Role and Function of the Secretory Metabolites Released from the Yeast.

PubMed

Ran, Chao; Huang, Lu; Liu, Zhi; Xu, Li; Yang, Yalin; Tacon, Philippe; Auclair, Eric; Zhou, Zhigang

2015-01-01

Yeast is frequently used as a probiotic in aquaculture with the potential to substitute for antibiotics. In this study, the involvement and extent to which the viability of yeast cells and thus the secretory metabolites released from the yeast contribute to effects of baker's yeast was investigated in Nile tilapia. No yeast, live yeast or heat-inactivated baker's yeast were added to basal diets high in fishmeal and low in soybean (diet A) or low in fishmeal and high in soybean (diet B), which were fed to fish for 8 weeks. Growth, feed utilization, gut microvilli morphology, and expressions of hsp70 and inflammation-related cytokines in the intestine and head kidney were assessed. Intestinal microbiota was investigated using 16S rRNA gene pyrosequencing. Gut alkaline phosphatase (AKP) activity was measured after challenging the fish with Aeromonas hydrophila. Results showed that live yeast significantly improved FBW and WG (P < 0.05), and tended to improve FCR (P = 0.06) of fish compared to the control (no yeast). No significant differences were observed between inactivated yeast and control. Live yeast improved gut microvilli length (P < 0.001) and density (P < 0.05) while inactivated yeast did not. The hsp70 expression level in both the intestine and head kidney of fish was significantly reduced by live yeast (P < 0.05) but not inactivated yeast. Live yeast but not inactivated yeast reduced intestinal expression of tnfα (P < 0.05), tgfβ (P < 0.05 under diet A) and il1β (P = 0.08). Intestinal Lactococcus spp. numbers were enriched by both live and inactivated yeast. Lastly, both live and inactivated yeast reduced the gut AKP activity compared to the control (P < 0.001), indicating protection of the host against infection by A. hydrophila. In conclusion, secretory metabolites did not play major roles in the growth promotion and disease protection effects of yeast. Nevertheless, secretory metabolites were the major contributing factor towards improved gut microvilli morphology, relieved stress status, and reduced intestinal inflammation of Nile tilapia fed diets supplemented with baker's yeast.

Extracellular Polysaccharides Produced by Yeasts and Yeast-Like Fungi

NASA Astrophysics Data System (ADS)

van Bogaert, Inge N. A.; de Maeseneire, Sofie L.; Vandamme, Erick J.

Several yeasts and yeast-like fungi are known to produce extracellular polysaccharides. Most of these contain D-mannose, either alone or in combination with other sugars or phosphate. A large chemical and structural variability is found between yeast species and even among different strains. The types of polymers that are synthesized can be chemically characterized as mannans, glucans, phosphoman-nans, galactomannans, glucomannans and glucuronoxylomannans. Despite these differences, almost all of the yeast exopolysaccharides display some sort of biological activity. Some of them have already applications in chemistry, pharmacy, cosmetics or as probiotic. Furthermore, some yeast exopolysaccharides, such as pullulan, exhibit specific physico-chemical and rheological properties, making them useful in a wide range of technical applications. A survey is given here of the production, the characteristics and the application potential of currently well studied yeast extracellular polysaccharides.

Solid-Body Fuse Developed for High- Voltage Space Power Missions

NASA Technical Reports Server (NTRS)

Dolce, James L.; Baez, Anastacio N.

2001-01-01

AEM Incorporated has completed the development, under a NASA Glenn Research Center contract, of a solid-body fuse for high-voltage power systems of satellites and spacecraft systems. High-reliability fuses presently defined by MIL-PRF-23419 do not meet the increased voltage and amperage requirements for the next generation of spacecraft. Solid-body fuses exhibit electrical and mechanical attributes that enable these fuses to perform reliably in the vacuum and high-vibration and -shock environments typically present in spacecraft applications. The construction and screening techniques for solid-body fuses described by MIL-PRF-23419/12 offer an excellent roadmap for the development of high-voltage solid-body fuses.

Studying p53 family proteins in yeast: Induction of autophagic cell death and modulation by interactors and small molecules

DOE Office of Scientific and Technical Information (OSTI.GOV)

Leão, Mariana; Gomes, Sara; Bessa, Cláudia

In this work, the yeast Saccharomyces cerevisiae was used to individually study human p53, p63 (full length and truncated forms) and p73. Using this cell system, the effect of these proteins on cell proliferation and death, and the influence of MDM2 and MDMX on their activities were analyzed. When expressed in yeast, wild-type p53, TAp63, ΔNp63 and TAp73 induced growth inhibition associated with S-phase cell cycle arrest. This growth inhibition was accompanied by reactive oxygen species production and autophagic cell death. Furthermore, they stimulated rapamycin-induced autophagy. On the contrary, none of the tested p53 family members induced apoptosis either permore » se or after apoptotic stimuli. As previously reported for p53, also TAp63, ΔNp63 and TAp73 increased actin expression levels and its depolarization, suggesting that ACT1 is also a p63 and p73 putative yeast target gene. Additionally, MDM2 and MDMX inhibited the activity of all tested p53 family members in yeast, although the effect was weaker on TAp63. Moreover, Nutlin-3a and SJ-172550 were identified as potential inhibitors of the p73 interaction with MDM2 and MDMX, respectively. Altogether, the yeast-based assays herein developed can be envisaged as a simplified cell system to study the involvement of p53 family members in autophagy, the modulation of their activities by specific interactors (MDM2 and MDMX), and the potential of new small molecules to modulate these interactions. - Highlights: • p53, p63 and p73 are individually studied in the yeast S. cerevisiae. • p53 family members induce ROS production, cell cycle arrest and autophagy in yeast. • p53 family members increase actin depolarization and expression levels in yeast. • MDM2 and MDMX inhibit the activity of p53 family members in yeast. • Yeast can be a useful tool to study the biology and drugability of p53, p63 and p73.« less

Phenotypic Diagnosis of Lineage and Differentiation During Sake Yeast Breeding

PubMed Central

Ohnuki, Shinsuke; Okada, Hiroki; Friedrich, Anne; Kanno, Yoichiro; Goshima, Tetsuya; Hasuda, Hirokazu; Inahashi, Masaaki; Okazaki, Naoto; Tamura, Hiroyasu; Nakamura, Ryo; Hirata, Dai; Fukuda, Hisashi; Shimoi, Hitoshi; Kitamoto, Katsuhiko; Watanabe, Daisuke; Schacherer, Joseph; Akao, Takeshi; Ohya, Yoshikazu

2017-01-01

Sake yeast was developed exclusively in Japan. Its diversification during breeding remains largely uncharacterized. To evaluate the breeding processes of the sake lineage, we thoroughly investigated the phenotypes and differentiation of 27 sake yeast strains using high-dimensional, single-cell, morphological phenotyping. Although the genetic diversity of the sake yeast lineage is relatively low, its morphological diversity has expanded substantially compared to that of the Saccharomyces cerevisiae species as a whole. Evaluation of the different types of breeding processes showed that the generation of hybrids (crossbreeding) has more profound effects on cell morphology than the isolation of mutants (mutation breeding). Analysis of phenotypic robustness revealed that some sake yeast strains are more morphologically heterogeneous, possibly due to impairment of cellular network hubs. This study provides a new perspective for studying yeast breeding genetics and micro-organism breeding strategies. PMID:28642365

Carbon source utilization and inhibitor tolerance of 45 oleaginous yeast species

PubMed Central

Sitepu, Irnayuli; Selby, Tylan; Lin, Ting; Zhu, Shirley; Boundy-Mills, Kyria

2014-01-01

Conversion of lignocellulosic hydrolysates to lipids using oleaginous (high lipid) yeasts requires alignment of the hydrolysate composition with the characteristics of the yeast strain, including ability to utilize certain nutrients, ability to grow independently of costly nutrients such as vitamins, and ability to tolerate inhibitors. Some combination of these characteristics may be present in wild strains. In this study, 48 oleaginous yeast strains belonging to 45 species were tested for ability to utilize carbon sources associated with lignocellulosic hydrolysates, tolerate inhibitors, and grow in medium without supplemented vitamins. Some well-studied oleaginous yeast species, as well as some that have not been frequently utilized in research or industrial production, emerged as promising candidates for industrial use due to ability to utilize many carbon sources, including Cryptococcus aureus, Cryptococcus laurentii, Hanaella aff. zeae, Tremella encephala, and Trichosporon coremiiforme. Other species excelled in inhibitor tolerance, including Candida aff. tropicalis, Cyberlindnera jadinii, Metschnikowia pulcherrima Schwanniomyces occidentalis and Wickerhamomyces ciferii. No yeast tested could utilize all carbon sources and tolerate all inhibitors tested. These results indicate that yeast strains should be selected based on characteristics compatible with the composition of the targeted hydrolysate. Other factors to consider include the production of valuable co-products such as carotenoids, availability of genetic tools, biosafety level, and flocculation of the yeast strain. The data generated in this study will aid in aligning yeasts with compatible hydrolysates for conversion of carbohydrates to lipids to be used for biofuels and other oleochemicals. PMID:24818698

Comparison of Three Commercial Systems for Identification of Yeasts Commonly Isolated in the Clinical Microbiology Laboratory

PubMed Central

Wadlin, Jill K.; Hanko, Gayle; Stewart, Rebecca; Pape, John; Nachamkin, Irving

1999-01-01

We evaluated three commercial systems (RapID Yeast Plus System; Innovative Diagnostic Systems, Norcross, Ga.; API 20C Aux; bioMerieux-Vitek, Hazelwood, Mo.; and Vitek Yeast Biochemical Card, bioMerieux-Vitek) against an auxinographic and microscopic morphologic reference method for the ability to identify yeasts commonly isolated in our clinical microbiology laboratory. Two-hundred one yeast isolates were compared in the study. The RapID Yeast Plus System was significantly better than either API 20C Aux (193 versus 167 correct identifications; P < 0.0001) or the Vitek Yeast Biochemical Card (193 versus 173 correct identifications; P = 0.003) for obtaining correct identifications to the species level without additional testing. There was no significant difference between results obtained with API 20C Aux and the Vitek Yeast Biochemical Card system (P = 0.39). The API 20C Aux system did not correctly identify any of the Candida krusei isolates (n = 23) without supplemental testing and accounted for the major differences between the API 20C Aux and RapID Yeast Plus systems. Overall, the RapID Yeast Plus System was easy to use and is a good system for the routine identification of clinically relevant yeasts. PMID:10325356

Indole-3-Acetic Acid-Producing Yeasts in the Phyllosphere of the Carnivorous Plant Drosera indica L

PubMed Central

Shin, Li-Ying; Wei, Jyuan-Yu; Fu, Shih-Feng; Chou, Jui-Yu

2014-01-01

Yeasts are widely distributed in nature and exist in association with other microorganisms as normal inhabitants of soil, vegetation, and aqueous environments. In this study, 12 yeast strains were enriched and isolated from leaf samples of the carnivorous plant Drosera indica L., which is currently threatened because of restricted habitats and use in herbal industries. According to similarities in large subunit and small subunit ribosomal RNA gene sequences, we identified 2 yeast species in 2 genera of the phylum Ascomycota, and 5 yeast species in 5 genera of the phylum Basidiomycota. All of the isolated yeasts produced indole-3-acetic acid (IAA) when cultivated in YPD broth supplemented with 0.1% L-tryptophan. Growth conditions, such as the pH and temperature of the medium, influenced yeast IAA production. Our results also suggested the existence of a tryptophan-independent IAA biosynthetic pathway. We evaluated the effects of various concentrations of exogenous IAA on yeast growth and observed that IAA produced by wild yeasts modifies auxin-inducible gene expression in Arabidopsis. Our data suggest that yeasts can promote plant growth and support ongoing prospecting of yeast strains for inclusion into biofertilizer for sustainable agriculture. PMID:25464336

Independent and additive effects of glutamic acid and methionine on yeast longevity.

PubMed

Wu, Ziyun; Song, Lixia; Liu, Shao Quan; Huang, Dejian

2013-01-01

It is established that glucose restriction extends yeast chronological and replicative lifespan, but little is known about the influence of amino acids on yeast lifespan, although some amino acids were reported to delay aging in rodents. Here we show that amino acid composition greatly alters yeast chronological lifespan. We found that non-essential amino acids (to yeast) methionine and glutamic acid had the most significant impact on yeast chronological lifespan extension, restriction of methionine and/or increase of glutamic acid led to longevity that was not the result of low acetic acid production and acidification in aging media. Remarkably, low methionine, high glutamic acid and glucose restriction additively and independently extended yeast lifespan, which could not be further extended by buffering the medium (pH 6.0). Our preliminary findings using yeasts with gene deletion demonstrate that glutamic acid addition, methionine and glucose restriction prompt yeast longevity through distinct mechanisms. This study may help to fill a gap in yeast model for the fast developing view that nutrient balance is a critical factor to extend lifespan.

Independent and Additive Effects of Glutamic Acid and Methionine on Yeast Longevity

PubMed Central

Wu, Ziyun; Song, Lixia; Liu, Shao Quan; Huang, Dejian

2013-01-01

It is established that glucose restriction extends yeast chronological and replicative lifespan, but little is known about the influence of amino acids on yeast lifespan, although some amino acids were reported to delay aging in rodents. Here we show that amino acid composition greatly alters yeast chronological lifespan. We found that non-essential amino acids (to yeast) methionine and glutamic acid had the most significant impact on yeast chronological lifespan extension, restriction of methionine and/or increase of glutamic acid led to longevity that was not the result of low acetic acid production and acidification in aging media. Remarkably, low methionine, high glutamic acid and glucose restriction additively and independently extended yeast lifespan, which could not be further extended by buffering the medium (pH 6.0). Our preliminary findings using yeasts with gene deletion demonstrate that glutamic acid addition, methionine and glucose restriction prompt yeast longevity through distinct mechanisms. This study may help to fill a gap in yeast model for the fast developing view that nutrient balance is a critical factor to extend lifespan. PMID:24244480

Genome dynamics and evolution in yeasts: A long-term yeast-bacteria competition experiment

PubMed Central

Katz, Michael; Knecht, Wolfgang; Compagno, Concetta; Piškur, Jure

2018-01-01

There is an enormous genetic diversity evident in modern yeasts, but our understanding of the ecological basis of such diversifications in nature remains at best fragmented so far. Here we report a long-term experiment mimicking a primordial competitive environment, in which yeast and bacteria co-exist and compete against each other. Eighteen yeasts covering a wide phylogenetic background spanning approximately 250 million years of evolutionary history were used to establish independent evolution lines for at most 130 passages. Our collection of hundreds of modified strains generated through such a rare two-species cross-kingdom competition experiment re-created the appearance of large-scale genomic rearrangements and altered phenotypes important in the diversification history of yeasts. At the same time, the methodology employed in this evolutionary study would also be a non-gene-technological method of reprogramming yeast genomes and then selecting yeast strains with desired traits. Cross-kingdom competition may therefore be a method of significant value to generate industrially useful yeast strains with new metabolic traits. PMID:29624585

Analysis of a genome-wide set of gene deletions in the fission yeast Schizosaccharomyces pombe

PubMed Central

Duhig, Trevor; Nam, Miyoung; Palmer, Georgia; Han, Sangjo; Jeffery, Linda; Baek, Seung-Tae; Lee, Hyemi; Shim, Young Sam; Lee, Minho; Kim, Lila; Heo, Kyung-Sun; Noh, Eun Joo; Lee, Ah-Reum; Jang, Young-Joo; Chung, Kyung-Sook; Choi, Shin-Jung; Park, Jo-Young; Park, Youngwoo; Kim, Hwan Mook; Park, Song-Kyu; Park, Hae-Joon; Kang, Eun-Jung; Kim, Hyong Bai; Kang, Hyun-Sam; Park, Hee-Moon; Kim, Kyunghoon; Song, Kiwon; Song, Kyung Bin; Nurse, Paul; Hoe, Kwang-Lae

2014-01-01

SUMMARY We report the construction and analysis of 4,836 heterozygous diploid deletion mutants covering 98.4% of the fission yeast genome. This resource provides a powerful tool for biotechnological and eukaryotic cell biology research. Comprehensive gene dispensability comparisons with budding yeast, the first time such studies have been possible between two eukaryotes, revealed that 83% of single copy orthologues in the two yeasts had conserved dispensability. Gene dispensability differed for certain pathways between the two yeasts, including mitochondrial translation and cell cycle checkpoint control. We show that fission yeast has more essential genes than budding yeast and that essential genes are more likely than non-essential genes to be single copy, broadly conserved and to contain introns. Growth fitness analyses determined sets of haploinsufficient and haploproficient genes for fission yeast, and comparisons with budding yeast identified specific ribosomal proteins and RNA polymerase subunits, which may act more generally to regulate eukaryotic cell growth. PMID:20473289

KSC-99pp0719

NASA Image and Video Library

1999-06-19

Workers in the launch tower at Launch Pad 17A, Cape Canaveral Air Station, help guide the first segment of the fairing around NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite. The satellite is scheduled for launch June 24 aboard a Boeing Delta II rocket. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study those elements to unlock the secrets of how galaxies evolve and to discover what the Universe was like when it was only a few minutes old

KSC-99pp0722

NASA Image and Video Library

1999-06-19

At Launch Pad 17A, Cape Canaveral Air Station, NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite (foreground) is partially covered by half of the fairing (behind it) that will protect it during launch. The satellite is scheduled for launch June 24 aboard a Boeing Delta II rocket. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study those elements to unlock the secrets of how galaxies evolve and to discover what the Universe was like when it was only a few minutes old

KSC-99pp0717

NASA Image and Video Library

1999-06-19

NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite sits ready for the fairing installation at Launch Pad 17A, Cape Canaveral Air Station. The satellite is scheduled for launch June 24 aboard a Boeing Delta II rocket. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe

Human gut Bacteroidetes can utilize yeast mannan through a selfish mechanism.

PubMed

Cuskin, Fiona; Lowe, Elisabeth C; Temple, Max J; Zhu, Yanping; Cameron, Elizabeth; Pudlo, Nicholas A; Porter, Nathan T; Urs, Karthik; Thompson, Andrew J; Cartmell, Alan; Rogowski, Artur; Hamilton, Brian S; Chen, Rui; Tolbert, Thomas J; Piens, Kathleen; Bracke, Debby; Vervecken, Wouter; Hakki, Zalihe; Speciale, Gaetano; Munōz-Munōz, Jose L; Day, Andrew; Peña, Maria J; McLean, Richard; Suits, Michael D; Boraston, Alisdair B; Atherly, Todd; Ziemer, Cherie J; Williams, Spencer J; Davies, Gideon J; Abbott, D Wade; Martens, Eric C; Gilbert, Harry J

2015-01-08

Yeasts, which have been a component of the human diet for at least 7,000 years, possess an elaborate cell wall α-mannan. The influence of yeast mannan on the ecology of the human microbiota is unknown. Here we show that yeast α-mannan is a viable food source for the Gram-negative bacterium Bacteroides thetaiotaomicron, a dominant member of the microbiota. Detailed biochemical analysis and targeted gene disruption studies support a model whereby limited cleavage of α-mannan on the surface generates large oligosaccharides that are subsequently depolymerized to mannose by the action of periplasmic enzymes. Co-culturing studies showed that metabolism of yeast mannan by B. thetaiotaomicron presents a 'selfish' model for the catabolism of this difficult to breakdown polysaccharide. Genomic comparison with B. thetaiotaomicron in conjunction with cell culture studies show that a cohort of highly successful members of the microbiota has evolved to consume sterically-restricted yeast glycans, an adaptation that may reflect the incorporation of eukaryotic microorganisms into the human diet.

Nectar yeasts: a natural microcosm for ecology.

PubMed

Chappell, Callie R; Fukami, Tadashi

2018-06-01

The species of yeasts that colonize floral nectar can modify the mutualistic relationships between plants and pollinators by changing the chemical properties of nectar. Recent evidence supporting this possibility has led to increased interest among ecologists in studying these fungi as well as the bacteria that interact with them in nectar. Although not fully explored, nectar yeasts also constitute a promising natural microcosm that can be used to facilitate development of general ecological theory. We discuss the methodological and conceptual advantages of using nectar yeasts from this perspective, including simplicity of communities, tractability of dispersal, replicability of community assembly, and the ease with which the mechanisms of species interactions can be studied in complementary experiments conducted in the field and the laboratory. To illustrate the power of nectar yeasts as a study system, we discuss several topics in community ecology, including environmental filtering, priority effects, and metacommunity dynamics. An exciting new direction is to integrate metagenomics and comparative genomics into nectar yeast research to address these fundamental ecological topics. Copyright © 2018 John Wiley & Sons, Ltd.

Micro-organisms behind the pollination scenes: microbial imprint on floral nectar sugar variation in a tropical plant community.

PubMed

Canto, A; Herrera, C M

2012-11-01

Variation in the composition of floral nectar reflects intrinsic plant characteristics as well as the action of extrinsic factors. Micro-organisms, particularly yeasts, represent one extrinsic factor that inhabit the nectar of animal-pollinated flowers worldwide. In this study a 'microbial imprint hypothesis' is formulated and tested, in which it is proposed that natural community-wide variation in nectar sugar composition will partly depend on the presence of yeasts in flowers. Occurrence and density of yeasts were studied microscopically in single-flower nectar samples of 22 animal-pollinated species from coastal xeric and sub-humid tropical habitats of the Yucatán Peninsula, Mexico. Nectar sugar concentration and composition were concurrently determined on the same samples using high-performance liquid chromatography (HPLC) methods. Microscopical examination of nectar samples revealed the presence of yeasts in nearly all plant species (21 out of 22 species) and in about half of the samples examined (51·8 % of total, all species combined). Plant species and individuals differed significantly in nectar sugar concentration and composition, and also in the incidence of nectar yeasts. After statistically controlling for differences between plant species and individuals, nectar yeasts still accounted for a significant fraction of community-wide variance in all nectar sugar parameters considered. Significant yeast × species interactions on sugar parameters revealed that plant species differed in the nectar sugar correlates of variation in yeast incidence. The results support the hypothesis that nectar yeasts impose a detectable imprint on community-wide variation in nectar sugar composition and concentration. Since nectar sugar features influence pollinator attraction and plant reproduction, future nectar studies should control for yeast presence and examine the extent to which microbial signatures on nectar characteristics ultimately have some influence on pollination services in plant communities.

Yeast community in traditional Portuguese Serpa cheese by culture-dependent and -independent DNA approaches.

PubMed

Gonçalves Dos Santos, Maria Teresa P; Benito, María José; Córdoba, María de Guía; Alvarenga, Nuno; Ruiz-Moyano Seco de Herrera, Santiago

2017-12-04

This study investigated the yeast community present in the traditional Portuguese cheese, Serpa, by culture-dependent and -independent methods. Sixteen batches of Serpa cheeses from various regional industries registered with the Protected Designation of Origin (PDO) versus non-PDO registered, during spring and winter, were used. Irrespective of the producer, the yeast counts were around 5log CFU/g in winter and, overall, were lower in spring. The yeast species identified at the end of ripening (30days), using PCR-RFLP analysis and sequencing of the 26S rRNA, mainly corresponded to Debaryomyces hansenii and Kluyveromyces marxianus, with Candida spp. and Pichia spp. present to a lesser extent. The culture-independent results, obtained using high-throughput sequencing analysis, confirmed the prevalence of Debaryomyces spp. and Kluyveromyces spp. but, also, that Galactomyces spp. was relevant for three of the five producers, which indicates its importance during the early stages of the cheese ripening process, considering it was not found among the dominant viable yeast species. In addition, differences between the identified yeast isolated from cheeses obtained from PDO and non-PDO registered industries, showed that the lack of regulation of the cheese-making practice, may unfavourably influence the final yeast microbiota. The new knowledge provided by this study of the yeast diversity in Serpa cheese, could be used to modify the cheese ripening conditions, to favour desirable yeast species. Additionally, the prevalent yeast isolates identified, Debaryomyces hansenii and Kluyveromyces spp., may have an important role during cheese ripening and in the final sensorial characteristics. Thus, the study of their technological and functional properties could be relevant, in the development of an autochthonous starter culture, to ensure final quality and safety of the cheese. Copyright © 2017 Elsevier B.V. All rights reserved.

Human COQ9 Rescues a coq9 Yeast Mutant by Enhancing Coenzyme Q Biosynthesis from 4-Hydroxybenzoic Acid and Stabilizing the CoQ-Synthome

PubMed Central

He, Cuiwen H.; Black, Dylan S.; Allan, Christopher M.; Meunier, Brigitte; Rahman, Shamima; Clarke, Catherine F.

2017-01-01

Coq9 is required for the stability of a mitochondrial multi-subunit complex, termed the CoQ-synthome, and the deamination step of Q intermediates that derive from para-aminobenzoic acid (pABA) in yeast. In human, mutations in the COQ9 gene cause neonatal-onset primary Q10 deficiency. In this study, we determined whether expression of human COQ9 could complement yeast coq9 point or null mutants. We found that expression of human COQ9 rescues the growth of the temperature-sensitive yeast mutant, coq9-ts19, on a non-fermentable carbon source and increases the content of Q6, by enhancing Q biosynthesis from 4-hydroxybenzoic acid (4HB). To study the mechanism for the rescue by human COQ9, we determined the steady-state levels of yeast Coq polypeptides in the mitochondria of the temperature-sensitive yeast coq9 mutant expressing human COQ9. We show that the expression of human COQ9 significantly increased steady-state levels of yeast Coq4, Coq6, Coq7, and Coq9 at permissive temperature. Human COQ9 polypeptide levels persisted at non-permissive temperature. A small amount of the human COQ9 co-purified with tagged Coq6, Coq6-CNAP, indicating that human COQ9 interacts with the yeast Q-biosynthetic complex. These findings suggest that human COQ9 rescues the yeast coq9 temperature-sensitive mutant by stabilizing the CoQ-synthome and increasing Q biosynthesis from 4HB. This finding provides a powerful approach to studying the function of human COQ9 using yeast as a model. PMID:28736527

Heavy metal removal by caustic-treated yeast immobilized in alginate

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu, Y.; Wilkins, E.

1995-12-31

Saccharomyces cerevisiae yeast biomass was treated with hot alkali to increase its biosorption capacity for heavy metals and then was immobilized in alginate gel. Biosorption capacities for Cu{sup 2+}, Cd{sup 2+}, and Zn{sup 2+} on alginate gel, native yeast, native yeast immobilized in alginate gel, and caustic-treated yeast immobilized in alginate gel were all compared. Immobilized yeasts could be reactivated and reused in a manner similar to the ion exchange resins. Immobilized caustic-treated yeast has high heavy metal biosorption capacity and high metal removal efficiency in a rather wide acidic pH region. The biosorption isotherm of immobilized caustic-treated yeast wasmore » studied, and empirical equations were obtained. The initial pH of polluted water affected the metal removal efficiency significantly, and the equilibrium biosorption capacity seemed to be temperature independent at lower initial metal concentrations.« less

METHOD OF SEPARATING FISSION PRODUCTS FROM FUSED BISMUTH-CONTAINING URANIUM

DOEpatents

Wiswall, R.H.

1958-06-24

A process is described for removing metal selectively from liquid metal compositions. The method effects separation of flssion product metals selectively from dilute solution in fused bismuth, which contains uraniunn in solution without removal of more than 1% of the uranium. The process comprises contacting the fused bismuth with a fused salt composition consisting of sodium, potassium and lithium chlorides, adding to fused bismuth and molten salt a quantity of bismuth chloride which is stoichiometrically required to convert the flssion product metals to be removed to their chlorides which are more stable in the fused salt than in the molten metal and are, therefore, preferentially taken up in the fused salt phase.

From drug to protein: using yeast genetics for high-throughput target discovery.

PubMed

Armour, Christopher D; Lum, Pek Yee

2005-02-01

The budding yeast Saccharomyces cerevisiae has long been an effective eukaryotic model system for understanding basic cellular processes. The genetic tractability and ease of manipulation in the laboratory make yeast well suited for large-scale chemical and genetic screens. Several recent studies describing the use of yeast genetics for high-throughput drug target identification are discussed in this review.

Dynamical analysis of yeast protein interaction network during the sake brewing process.

PubMed

Mirzarezaee, Mitra; Sadeghi, Mehdi; Araabi, Babak N

2011-12-01

Proteins interact with each other for performing essential functions of an organism. They change partners to get involved in various processes at different times or locations. Studying variations of protein interactions within a specific process would help better understand the dynamic features of the protein interactions and their functions. We studied the protein interaction network of Saccharomyces cerevisiae (yeast) during the brewing of Japanese sake. In this process, yeast cells are exposed to several stresses. Analysis of protein interaction networks of yeast during this process helps to understand how protein interactions of yeast change during the sake brewing process. We used gene expression profiles of yeast cells for this purpose. Results of our experiments revealed some characteristics and behaviors of yeast hubs and non-hubs and their dynamical changes during the brewing process. We found that just a small portion of the proteins (12.8 to 21.6%) is responsible for the functional changes of the proteins in the sake brewing process. The changes in the number of edges and hubs of the yeast protein interaction networks increase in the first stages of the process and it then decreases at the final stages.

Dietary live yeast alters metabolic profiles, protein biosynthesis and thermal stress tolerance of Drosophila melanogaster.

PubMed

Colinet, Hervé; Renault, David

2014-04-01

The impact of nutritional factors on insect's life-history traits such as reproduction and lifespan has been excessively examined; however, nutritional determinant of insect's thermal tolerance has not received a lot of attention. Dietary live yeast represents a prominent source of proteins and amino acids for laboratory-reared drosophilids. In this study, Drosophila melanogaster adults were fed on diets supplemented or not with live yeast. We hypothesized that manipulating nutritional conditions through live yeast supplementation would translate into altered physiology and stress tolerance. We verified how live yeast supplementation affected body mass characteristics, total lipids and proteins, metabolic profiles and cold tolerance (acute and chronic stress). Females fed with live yeast had increased body mass and contained more lipids and proteins. Using GC/MS profiling, we found distinct metabolic fingerprints according to nutritional conditions. Metabolite pathway enrichment analysis corroborated that live yeast supplementation was associated with amino acid and protein biosyntheses. The cold assays revealed that the presence of dietary live yeast greatly promoted cold tolerance. Hence, this study conclusively demonstrates a significant interaction between nutritional conditions and thermal tolerance. Copyright © 2014 Elsevier Inc. All rights reserved.

The impact of yeast fermentation on dough matrix properties.

PubMed

Rezaei, Mohammad N; Jayaram, Vinay B; Verstrepen, Kevin J; Courtin, Christophe M

2016-08-01

Most studies on dough properties are performed on yeastless dough to exclude the complicating, time-dependent effect of yeast. Baker's yeast, however, impacts dough matrix properties during fermentation, probably through the production of primary (CO2 and ethanol) and secondary (glycerol, acetic acid and succinic acid) metabolites. The aim of this study is to obtain a better understanding of the changes in yeasted dough behavior introduced by fermentation, by investigating the impact of yeast fermentation on Farinograph dough consistency, dough spread, Kieffer rig dough extensibility and gluten agglomeration behavior in a fermented dough-batter gluten starch separation system. Results show that fermentation leads to a dough with less flow and lower extensibility that breaks more easily under stress and strain. The dough showed less elastic and more plastic deformation behavior. Gluten agglomerates were smaller for yeasted dough than for the unyeasted control. These changes probably have to be attributed to metabolites generated during fermentation. Indeed, organic acids and also ethanol in concentrations produced by yeast were previously shown to have similar effects in yeastless dough. These findings imply the high importance of yeast fermentation metabolites on dough matrix properties in industrial bread production. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

Effects of wet etch processing on laser-induced damage of fused silica surfaces

DOE Office of Scientific and Technical Information (OSTI.GOV)

Battersby, C.L.; Kozlowski, M.R.; Sheehan, L.M.

1998-12-22

Laser-induced damage of transparent fused silica optical components by 355 nm illumination occurs primarily at surface defects produced during the grinding and polishing processes. These defects can either be surface defects or sub-surface damage.Wet etch processing in a buffered hydrogen fluoride (HF) solution has been examined as a tool for characterizing such defects. A study was conducted to understand the effects of etch depth on the damage threshold of fused silica substrates. The study used a 355 nm, 7.5 ns, 10 Hz Nd:YAG laser to damage test fused silica optics through various wet etch processing steps. Inspection of the surfacemore » quality was performed with Nomarski microscopy and Total Internal Reflection Microscopy. The damage test data and inspection results were correlated with polishing process specifics. The results show that a wet etch exposes subsurface damage while maintaining or improving the laser damage performance. The benefits of a wet etch must be evaluated for each polishing process.« less

Modifying Expression Modes of Human Neurotensin Receptor Type 1 Alters Sensing Capabilities for Agonists in Yeast Signaling Biosensor.

PubMed

Hashi, Hiroki; Nakamura, Yasuyuki; Ishii, Jun; Kondo, Akihiko

2018-04-01

Neurotensin receptor type 1 (NTSR1), a member of the G-protein-coupled receptor (GPCR) family, is naturally activated by binding of a neurotensin peptide, leading to a variety of physiological effects. The budding yeast Saccharomyces cerevisiae is a proven host organism for assaying the agonistic activation of human GPCRs. Previous studies showed that yeast cells can functionally express human NTSR1 receptor, permitting the detection of neurotensin-promoted signaling using a ZsGreen fluorescent reporter gene. However, the fluorescence intensity (sensitivity) of NTSR1-expressing yeast cells is low compared to that of yeast cells expressing other human GPCRs (e.g., human somatostatin receptors). The present study sought to increase the sensitivity of the NTSR1-expressing yeast for use as a fluorescent biosensor, including modification of the expression of human NTSR1 in yeast. Changes in the transcription, translation, and transport of the receptor are attempted by altering the promoter, consensus Kozak-like sequence, and secretion signal sequences of the NTSR1-encoding gene. The resulting yeast cells exhibited increased sensitivity to exogenously added peptide. The cells are further engineered by using cell-surface display technology to ensure that the agonistic peptides are secreted and tethered to the yeast cell wall, yielding cells with enhanced NTSR1 activation. This yeast biosensor holds promise for the identification of agonists to treat NTSR1-related diseases. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Feasibility of full-spectrum endoscopy: Korea’s first full-spectrum endoscopy colonoscopic trial

PubMed Central

Song, Jeong-Yeop; Cho, Youn Hee; Kim, Mi A; Kim, Jeong-Ae; Lee, Chun Tek; Lee, Moon Sung

2016-01-01

AIM: To evaluate the full-spectrum endoscopy (FUSE) colonoscopy system as the first report on the utility thereof in a Korean population. METHODS: We explored the efficacy of the FUSE colonoscopy in a retrospective, single-center feasibility study performed between February 1 and July 20, 2015. A total of 262 subjects (age range: 22-80) underwent the FUSE colonoscopy for colorectal cancer screening, polyp surveillance, or diagnostic evaluation. The cecal intubation success rate, the polyp detection rate (PDR), the adenoma detection rate (ADR), and the diverticulum detection rate (DDR), were calculated. Also, the success rates of therapeutic interventions were evaluated with biopsy confirmation. RESULTS: All patients completed the study and the success rates of cecal and terminal ileal intubation were 100% with the FUSE colonoscope; we found 313 polyps in 142 patients and 173 adenomas in 95. The overall PDR, ADR and DDR were 54.2%, 36.3%, and 25.2%, respectively, and were higher in males, and increased with age. The endoscopists and nurses involved considered that the full-spectrum colonoscope improved navigation and orientation within the colon. No colonoscopy was aborted because of colonoscope malfunction. CONCLUSION: The FUSE colonoscopy yielded a higher PDR, ADR, DDR than did traditional colonoscopy, without therapeutic failure or complications, showing feasible, effective, and safe in this first Korean trial. PMID:26937150

Metabolic Engineering of Oleaginous Yeasts for Production of Fuels and Chemicals.

PubMed

Shi, Shuobo; Zhao, Huimin

2017-01-01

Oleaginous yeasts have been increasingly explored for production of chemicals and fuels via metabolic engineering. Particularly, there is a growing interest in using oleaginous yeasts for the synthesis of lipid-related products due to their high lipogenesis capability, robustness, and ability to utilize a variety of substrates. Most of the metabolic engineering studies in oleaginous yeasts focused on Yarrowia that already has plenty of genetic engineering tools. However, recent advances in systems biology and synthetic biology have provided new strategies and tools to engineer those oleaginous yeasts that have naturally high lipid accumulation but lack genetic tools, such as Rhodosporidium , Trichosporon , and Lipomyces . This review highlights recent accomplishments in metabolic engineering of oleaginous yeasts and recent advances in the development of genetic engineering tools in oleaginous yeasts within the last 3 years.

Identification of a rice metal tolerance protein OsMTP11 as a manganese transporter

PubMed Central

Zhang, Mei; Liu, Baoxiu

2017-01-01

Metal tolerance proteins (MTPs) are a gene family of cation efflux transporters that occur widely in plants and might serve an essential role in metal homeostasis and tolerance. Our research describes the identification, characterization, and localization of OsMTP11, a member of the MTP family from rice. OsMTP11 was expressed constitutively and universally in different tissues in rice plant. Heterologous expression in yeast showed that OsMTP11 complemented the hypersensitivity of mutant strains to Mn, and also complemented yeast mutants to other metals, including Co and Ni. Real time RT-PCR analysis demonstrated OsMTP11 expression was substantially enhanced following 4 h under Cd, Zn, Ni, and Mn treatments, suggesting possible roles of OsMTP11 involvement in heavy metal stress responses. Promoter analysis by transgenic assays with GUS as a reporter gene and mRNA in situ hybridization experiments showed that OsMTP11 was expressed specifically in conducting tissues in rice. DNA methylation assays of genomic DNA in rice treated with Cd, Zn, Ni, and Mn revealed that decreased DNA methylation levels were present in the OsMTP11 promoter region, which was consistent with OsMTP11 induced-expression patterns resulting from heavy metal stress. This result suggested that DNA methylation is one of major factors regulating expression of OsMTP11 through epigenetic mechanisms. OsMTP11 fused to green fluorescent protein (GFP) localized to the entire onion epidermal cell cytoplasm, while vacuolar membrane exhibited increased GFP signals, consistent with an OsMTP11 function in cation sequestration. Our results indicated that OsMTP11 might play vital roles in Mn and other heavy metal transportation in rice. PMID:28394944

Crystal structure, mutational analysis and RNA-dependent ATPase activity of the yeast DEAD-box pre-mRNA splicing factor Prp28

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jacewicz, Agata; Schwer, Beate; Smith, Paul

Yeast Prp28 is a DEAD-box pre-mRNA splicing factor implicated in displacing U1 snRNP from the 5' splice site. Here we report that the 588-aa Prp28 protein consists of a trypsin-sensitive 126-aa N-terminal segment (of which aa 1–89 are dispensable for Prp28 function in vivo) fused to a trypsin-resistant C-terminal catalytic domain. Purified recombinant Prp28 and Prp28-(127–588) have an intrinsic RNA-dependent ATPase activity, albeit with a low turnover number. The crystal structure of Prp28-(127–588) comprises two RecA-like domains splayed widely apart. AMPPNP•Mg 2+ is engaged by the proximal domain, with proper and specific contacts from Phe194 and Gln201 (Q motif) tomore » the adenine nucleobase. The triphosphate moiety of AMPPNP•Mg 2+ is not poised for catalysis in the open domain conformation. Guided by the Prp28•AMPPNP structure, and that of the Drosophila Vasa•AMPPNP•Mg 2+•RNA complex, we targeted 20 positions in Prp28 for alanine scanning. ATP-site components Asp341 and Glu342 (motif II) and Arg527 and Arg530 (motif VI) and RNA-site constituent Arg476 (motif Va) are essential for Prp28 activity in vivo. Synthetic lethality of double-alanine mutations highlighted functionally redundant contacts in the ATP-binding (Phe194-Gln201, Gln201-Asp502) and RNA-binding (Arg264-Arg320) sites. As a result, overexpression of defective ATP-site mutants, but not defective RNA-site mutants, elicited severe dominant-negative growth defects.« less

Comparative analysis of programmed cell death pathways in filamentous fungi.

PubMed

Fedorova, Natalie D; Badger, Jonathan H; Robson, Geoff D; Wortman, Jennifer R; Nierman, William C

2005-12-08

Fungi can undergo autophagic- or apoptotic-type programmed cell death (PCD) on exposure to antifungal agents, developmental signals, and stress factors. Filamentous fungi can also exhibit a form of cell death called heterokaryon incompatibility (HI) triggered by fusion between two genetically incompatible individuals. With the availability of recently sequenced genomes of Aspergillus fumigatus and several related species, we were able to define putative components of fungi-specific death pathways and the ancestral core apoptotic machinery shared by all fungi and metazoa. Phylogenetic profiling of HI-associated proteins from four Aspergilli and seven other fungal species revealed lineage-specific protein families, orphan genes, and core genes conserved across all fungi and metazoa. The Aspergilli-specific domain architectures include NACHT family NTPases, which may function as key integrators of stress and nutrient availability signals. They are often found fused to putative effector domains such as Pfs, SesB/LipA, and a newly identified domain, HET-s/LopB. Many putative HI inducers and mediators are specific to filamentous fungi and not found in unicellular yeasts. In addition to their role in HI, several of them appear to be involved in regulation of cell cycle, development and sexual differentiation. Finally, the Aspergilli possess many putative downstream components of the mammalian apoptotic machinery including several proteins not found in the model yeast, Saccharomyces cerevisiae. Our analysis identified more than 100 putative PCD associated genes in the Aspergilli, which may help expand the range of currently available treatments for aspergillosis and other invasive fungal diseases. The list includes species-specific protein families as well as conserved core components of the ancestral PCD machinery shared by fungi and metazoa.

Identification of novel transcriptional regulators of Zat12 using comprehensive yeast one-hybrid screens.

PubMed

Ben Daniel, Bat-Hen; Cattan, Esther; Wachtel, Chaim; Avrahami, Dorit; Glick, Yair; Malichy, Asaf; Gerber, Doron; Miller, Gad

2016-08-01

To appropriately acclimate to environmental stresses, plants have to rapidly activate a specific transcriptional program. Yet, the identity and function of many of the transcriptional regulators that mediate early responses to abiotic stress stimuli is still unknown. In this work we employed the promoter of the multi-stress-responsive zinc-finger protein Zat12 in yeast one-hybrid (Y1H) screens to identify early abiotic stress-responsive transcriptional regulators. Analysis of Zat12 promoter fragments fused to luciferase underlined an approximately 200 bp fragment responsive to NaCl and to reactive oxygen species (ROS). Using these segments and others as baits against Y1H control or stress Arabidopsis prey libraries, we identified 15 potential Zat12 transcriptional regulators. Among the prominent proteins identified were known transcription factors including bZIP29 and ANAC91 as well as unknown function proteins such as a homolog of the human USB1, a U6 small nuclear RNA (snRNA) processing protein, and dormancy/auxin-associated family protein 2 (DRM2). Altered expression of Zat12 during high light stress in the knockout mutants further indicated the involvement of these proteins in the regulation of Zat12. Using a state of the art microfluidic approach we showed that AtUSB1 and DRM2 can specifically bind dsDNA and were able to identify the preferred DNA-binding motif of all four proteins. Overall, the proteins identified in this work provide an important start point for charting the earliest signaling network of Zat12 and of other genes required for acclimation to abiotic stresses. © 2016 Scandinavian Plant Physiology Society.

Amino-terminal enhancer of split (AES) interacts with the oncoprotein NUP98-HOXA9 and enhances its transforming ability.

PubMed

Sarma, Nayan J; Yaseen, Nabeel R

2011-11-11

NUP98-HOXA9 is the prototype of NUP98 fusion oncoproteins that cause acute myeloid leukemia. It consists of an N-terminal FG-rich portion of the nucleoporin NUP98 fused to the homeodomain region of the homeobox protein HOXA9, and acts as an aberrant transcription factor. To identify interacting partners of NUP98-HOXA9, we used a cytoplasmic yeast two-hybrid assay to avoid the nonspecific trans-activation that would occur with the traditional yeast two-hybrid assay due to the transactivating properties of NUP98-HOXA9. We identified amino-terminal enhancer of split (AES), a transcriptional regulator of the transducin-like enhancer/Groucho family as a novel interaction partner of NUP98-HOXA9. The interaction was confirmed by in vitro pulldown and co-immunoprecipitation assays and was shown to require the FG repeat region of NUP98-HOXA9. Immunofluorescence analysis showed that AES localizes primarily to the interior of the nucleus. AES also showed a strong interaction with wild-type NUP98. AES augmented the transcriptional activity of NUP98-HOXA9. In the presence of NUP98-HOXA9, AES caused an increase in long-term proliferation of primary human CD34+ cells with a marked increase in the numbers of primitive cells. These effects of AES were not observed in the absence of NUP98-HOXA9. AES knockdown diminished the transcriptional and proliferative effects of NUP98-HOXA9. AES caused a shift away from the erythroid lineage in cells expressing NUP98-HOXA9. These data establish AES as an interacting partner of NUP98-HOXA9 and show that it cooperates with NUP98-HOXA9 in transcriptional regulation and cell transformation.

Amino-terminal Enhancer of Split (AES) Interacts with the Oncoprotein NUP98-HOXA9 and Enhances Its Transforming Ability*

PubMed Central

Sarma, Nayan J.; Yaseen, Nabeel R.

2011-01-01

NUP98-HOXA9 is the prototype of NUP98 fusion oncoproteins that cause acute myeloid leukemia. It consists of an N-terminal FG-rich portion of the nucleoporin NUP98 fused to the homeodomain region of the homeobox protein HOXA9, and acts as an aberrant transcription factor. To identify interacting partners of NUP98-HOXA9, we used a cytoplasmic yeast two-hybrid assay to avoid the nonspecific trans-activation that would occur with the traditional yeast two-hybrid assay due to the transactivating properties of NUP98-HOXA9. We identified amino-terminal enhancer of split (AES), a transcriptional regulator of the transducin-like enhancer/Groucho family as a novel interaction partner of NUP98-HOXA9. The interaction was confirmed by in vitro pulldown and co-immunoprecipitation assays and was shown to require the FG repeat region of NUP98-HOXA9. Immunofluorescence analysis showed that AES localizes primarily to the interior of the nucleus. AES also showed a strong interaction with wild-type NUP98. AES augmented the transcriptional activity of NUP98-HOXA9. In the presence of NUP98-HOXA9, AES caused an increase in long-term proliferation of primary human CD34+ cells with a marked increase in the numbers of primitive cells. These effects of AES were not observed in the absence of NUP98-HOXA9. AES knockdown diminished the transcriptional and proliferative effects of NUP98-HOXA9. AES caused a shift away from the erythroid lineage in cells expressing NUP98-HOXA9. These data establish AES as an interacting partner of NUP98-HOXA9 and show that it cooperates with NUP98-HOXA9 in transcriptional regulation and cell transformation. PMID:21937451

Crystal structure, mutational analysis and RNA-dependent ATPase activity of the yeast DEAD-box pre-mRNA splicing factor Prp28

DOE PAGES

Jacewicz, Agata; Schwer, Beate; Smith, Paul; ...

2014-10-10

Yeast Prp28 is a DEAD-box pre-mRNA splicing factor implicated in displacing U1 snRNP from the 5' splice site. Here we report that the 588-aa Prp28 protein consists of a trypsin-sensitive 126-aa N-terminal segment (of which aa 1–89 are dispensable for Prp28 function in vivo) fused to a trypsin-resistant C-terminal catalytic domain. Purified recombinant Prp28 and Prp28-(127–588) have an intrinsic RNA-dependent ATPase activity, albeit with a low turnover number. The crystal structure of Prp28-(127–588) comprises two RecA-like domains splayed widely apart. AMPPNP•Mg 2+ is engaged by the proximal domain, with proper and specific contacts from Phe194 and Gln201 (Q motif) tomore » the adenine nucleobase. The triphosphate moiety of AMPPNP•Mg 2+ is not poised for catalysis in the open domain conformation. Guided by the Prp28•AMPPNP structure, and that of the Drosophila Vasa•AMPPNP•Mg 2+•RNA complex, we targeted 20 positions in Prp28 for alanine scanning. ATP-site components Asp341 and Glu342 (motif II) and Arg527 and Arg530 (motif VI) and RNA-site constituent Arg476 (motif Va) are essential for Prp28 activity in vivo. Synthetic lethality of double-alanine mutations highlighted functionally redundant contacts in the ATP-binding (Phe194-Gln201, Gln201-Asp502) and RNA-binding (Arg264-Arg320) sites. As a result, overexpression of defective ATP-site mutants, but not defective RNA-site mutants, elicited severe dominant-negative growth defects.« less

Exploring the chemodiversity and biological activities of the secondary metabolites from the marine fungus Neosartorya pseudofischeri.

PubMed

Liang, Wan-Ling; Le, Xiu; Li, Hou-Jin; Yang, Xiang-Ling; Chen, Jun-Xiong; Xu, Jun; Liu, Huan-Liang; Wang, Lai-You; Wang, Kun-Teng; Hu, Kun-Chao; Yang, De-Po; Lan, Wen-Jian

2014-11-24

The production of fungal metabolites can be remarkably influenced by various cultivation parameters. To explore the biosynthetic potentials of the marine fungus, Neosartorya pseudofischeri, which was isolated from the inner tissue of starfish Acanthaster planci, glycerol-peptone-yeast extract (GlyPY) and glucose-peptone-yeast extract (GluPY) media were used to culture this fungus. When cultured in GlyPY medium, this fungus produced two novel diketopiperazines, neosartins A and B (1 and 2), together with six biogenetically-related known diketopiperazines,1,2,3,4-tetrahydro-2, 3-dimethyl-1,4-dioxopyrazino[1,2-a]indole (3), 1,2,3,4-tetrahydro-2-methyl-3-methylen e-1,4-dioxopyrazino[1,2-a]indole (4), 1,2,3,4-tetrahydro-2-methyl-1,3,4-trioxopyrazino[1,2-a] indole (5), 6-acetylbis(methylthio)gliotoxin (10), bisdethiobis(methylthio)gliotoxin (11), didehydrobisdethiobis(methylthio)gliotoxin (12) and N-methyl-1H-indole-2-carboxamide (6). However, a novel tetracyclic-fused alkaloid, neosartin C (14), a meroterpenoid, pyripyropene A (15), gliotoxin (7) and five known gliotoxin analogues, acetylgliotoxin (8), reduced gliotoxin (9), 6-acetylbis(methylthio)gliotoxin (10), bisdethiobis(methylthio) gliotoxin (11) and bis-N-norgliovictin (13), were obtained when grown in glucose-containing medium (GluPY medium). This is the first report of compounds 3, 4, 6, 9, 10 and 12 as naturally occurring. Their structures were determined mainly by MS, 1D and 2D NMR data. The possible biosynthetic pathways of gliotoxin-related analogues and neosartin C were proposed. The antibacterial activity of compounds 2-14 and the cytotoxic activity of compounds 4, 5 and 7-13 were evaluated. Their structure-activity relationships are also preliminarily discussed.

A Rab5 GTPase module is important for autophagosome closure

PubMed Central

Lipatova, Zhanna; Sun, Dan; Zhu, Xiaolong; Li, Rui; Wu, Zulin; You, Weiming; Cong, Xiaoxia; Zhou, Yiting; Gyurkovska, Valeriya; Liu, Yutao; Li, Qunli; Li, Wenjing; Cheng, Jie; Segev, Nava

2017-01-01

In the conserved autophagy pathway, the double-membrane autophagosome (AP) engulfs cellular components to be delivered for degradation in the lysosome. While only sealed AP can productively fuse with the lysosome, the molecular mechanism of AP closure is currently unknown. Rab GTPases, which regulate all intracellular trafficking pathways in eukaryotes, also regulate autophagy. Rabs function in GTPase modules together with their activators and downstream effectors. In yeast, an autophagy-specific Ypt1 GTPase module, together with a set of autophagy-related proteins (Atgs) and a phosphatidylinositol-3-phosphate (PI3P) kinase, regulates AP formation. Fusion of APs and endosomes with the vacuole (the yeast lysosome) requires the Ypt7 GTPase module. We have previously shown that the Rab5-related Vps21, within its endocytic GTPase module, regulates autophagy. However, it was not clear which autophagy step it regulates. Here, we show that this module, which includes the Vps9 activator, the Rab5-related Vps21, the CORVET tethering complex, and the Pep12 SNARE, functions after AP expansion and before AP closure. Whereas APs are not formed in mutant cells depleted for Atgs, sealed APs accumulate in cells depleted for the Ypt7 GTPase module members. Importantly, depletion of individual members of the Vps21 module results in a novel phenotype: accumulation of unsealed APs. In addition, we show that Vps21-regulated AP closure precedes another AP maturation step, the previously reported PI3P phosphatase-dependent Atg dissociation. Our results delineate three successive steps in the autophagy pathway regulated by Rabs, Ypt1, Vps21 and Ypt7, and provide the first insight into the upstream regulation of AP closure. PMID:28934205

Oral yeast colonization throughout pregnancy

PubMed Central

Rio, Rute; Simões-Silva, Liliana; Garro, Sofia; Silva, Mário-Jorge; Azevedo, Álvaro

2017-01-01

Background Recent studies suggest that placenta may harbour a unique microbiome that may have origin in maternal oral microbiome. Although the major physiological and hormonal adjustments observed in pregnant women lead to biochemical and microbiological modifications of the oral environment, very few studies evaluated the changes suffered by the oral microbiota throughout pregnancy. So, the aim of our study was to evaluate oral yeast colonization throughout pregnancy and to compare it with non-pregnant women. Material and Methods The oral yeast colonization was assessed in saliva of 30 pregnant and non-pregnant women longitudinally over a 6-months period. Demographic information was collected, a non-invasive intra-oral examination was performed and saliva flow and pH were determined. Results Pregnant and non-pregnant groups were similar regarding age and level of education. Saliva flow rate did not differ, but saliva pH was lower in pregnant than in non-pregnant women. Oral yeast prevalence was higher in pregnant than in non-pregnant women, either in the first or in the third trimester, but did not attain statistical significance. In individuals colonized with yeast, the total yeast quantification (Log10CFU/mL) increase from the 1st to the 3rd trimester in pregnant women, but not in non-pregnant women. Conclusions Pregnancy may favour oral yeast growth that may be associated with an acidic oral environment. Key words:Oral yeast, fungi, pregnancy, saliva pH. PMID:28160578

Fermentation behaviour and volatile compound production by agave and grape must yeasts in high sugar Agave tequilana and grape must fermentations.

PubMed

Arrizon, Javier; Fiore, Concetta; Acosta, Guillermina; Romano, Patrizia; Gschaedler, Anne

2006-01-01

Few studies have been performed on the characterization of yeasts involved in the production of agave distilled beverages and their individual fermentation properties. In this study, a comparison and evaluation of yeasts of different origins in the tequila and wine industries were carried out for technological traits. Fermentations were carried out in high (300 g l(-1)) and low (30 g l(-1)) sugar concentrations of Agave tequilana juice, in musts obtained from Fiano (white) and Aglianico (red) grapes and in YPD medium (with 270 g l(-1) of glucose added) as a control. Grape yeasts exhibited a reduced performance in high-sugar agave fermentation, while both agave and grape yeasts showed similar fermentation behaviour in grape musts. Production levels of volatile compounds by grape and agave yeasts differed in both fermentations.

Human gut Bacteroidetes can utilize yeast mannan through a selfish mechanism

PubMed Central

Zhu, Yanping; Cameron, Elizabeth; Pudlo, Nicholas A.; Porter, Nathan T.; Urs, Karthik; Thompson, Andrew J.; Cartmell, Alan; Rogowski, Artur; Hamilton, Brian S.; Chen, Rui; Tolbert, Thomas J.; Piens, Kathleen; Bracke, Debby; Vervecken, Wouter; Hakki, Zalihe; Speciale, Gaetano; Munōz-Munōz, Jose L.; Day, Andrew; Peña, Maria J.; McLean, Richard; Suits, Michael D.; Boraston, Alisdair B.; Atherly, Todd; Ziemer, Cherie J.; Williams, Spencer J.; Davies, Gideon J.; Abbott, D. Wade; Martens, Eric C.; Gilbert, Harry J.

2016-01-01

Yeasts, which have been a component of the human diet for at least 7000 years, possess an elaborate cell wall α-mannan. The influence of yeast mannan on the ecology of the human microbiota is unknown. Here we show that yeast α-mannan is a viable food source for Bacteroides thetaiotaomicron (Bt), a dominant member of the microbiota. Detailed biochemical analysis and targeted gene disruption studies support a model whereby limited cleavage of α-mannan on the surface generates large oligosaccharides that are subsequently depolymerized to mannose by the action of periplasmic enzymes. Co-culturing studies showed that metabolism of yeast mannan by Bt presents a ‘selfish’ model for the catabolism of this recalcitrant polysaccharide. This report shows how a cohort of highly successful members of the microbiota has evolved to consume sterically-restricted yeast glycans, an adaptation that may reflect the incorporation of eukaryotic microorganisms into the human diet. PMID:25567280

Budding yeast for budding geneticists: a primer on the Saccharomyces cerevisiae model system.

PubMed

Duina, Andrea A; Miller, Mary E; Keeney, Jill B

2014-05-01

The budding yeast Saccharomyces cerevisiae is a powerful model organism for studying fundamental aspects of eukaryotic cell biology. This Primer article presents a brief historical perspective on the emergence of this organism as a premier experimental system over the course of the past century. An overview of the central features of the S. cerevisiae genome, including the nature of its genetic elements and general organization, is also provided. Some of the most common experimental tools and resources available to yeast geneticists are presented in a way designed to engage and challenge undergraduate and graduate students eager to learn more about the experimental amenability of budding yeast. Finally, a discussion of several major discoveries derived from yeast studies highlights the far-reaching impact that the yeast system has had and will continue to have on our understanding of a variety of cellular processes relevant to all eukaryotes, including humans.

Budding Yeast for Budding Geneticists: A Primer on the Saccharomyces cerevisiae Model System

PubMed Central

Duina, Andrea A.; Miller, Mary E.; Keeney, Jill B.

2014-01-01

The budding yeast Saccharomyces cerevisiae is a powerful model organism for studying fundamental aspects of eukaryotic cell biology. This Primer article presents a brief historical perspective on the emergence of this organism as a premier experimental system over the course of the past century. An overview of the central features of the S. cerevisiae genome, including the nature of its genetic elements and general organization, is also provided. Some of the most common experimental tools and resources available to yeast geneticists are presented in a way designed to engage and challenge undergraduate and graduate students eager to learn more about the experimental amenability of budding yeast. Finally, a discussion of several major discoveries derived from yeast studies highlights the far-reaching impact that the yeast system has had and will continue to have on our understanding of a variety of cellular processes relevant to all eukaryotes, including humans. PMID:24807111

A Comparison of the Beneficial Effects of Live and Heat-Inactivated Baker’s Yeast on Nile Tilapia: Suggestions on the Role and Function of the Secretory Metabolites Released from the Yeast

PubMed Central

Liu, Zhi; Xu, Li; Yang, Yalin; Tacon, Philippe; Auclair, Eric; Zhou, Zhigang

2015-01-01

Yeast is frequently used as a probiotic in aquaculture with the potential to substitute for antibiotics. In this study, the involvement and extent to which the viability of yeast cells and thus the secretory metabolites released from the yeast contribute to effects of baker’s yeast was investigated in Nile tilapia. No yeast, live yeast or heat-inactivated baker’s yeast were added to basal diets high in fishmeal and low in soybean (diet A) or low in fishmeal and high in soybean (diet B), which were fed to fish for 8 weeks. Growth, feed utilization, gut microvilli morphology, and expressions of hsp70 and inflammation-related cytokines in the intestine and head kidney were assessed. Intestinal microbiota was investigated using 16S rRNA gene pyrosequencing. Gut alkaline phosphatase (AKP) activity was measured after challenging the fish with Aeromonas hydrophila. Results showed that live yeast significantly improved FBW and WG (P < 0.05), and tended to improve FCR (P = 0.06) of fish compared to the control (no yeast). No significant differences were observed between inactivated yeast and control. Live yeast improved gut microvilli length (P < 0.001) and density (P < 0.05) while inactivated yeast did not. The hsp70 expression level in both the intestine and head kidney of fish was significantly reduced by live yeast (P < 0.05) but not inactivated yeast. Live yeast but not inactivated yeast reduced intestinal expression of tnfα (P < 0.05), tgfβ (P < 0.05 under diet A) and il1β (P = 0.08). Intestinal Lactococcus spp. numbers were enriched by both live and inactivated yeast. Lastly, both live and inactivated yeast reduced the gut AKP activity compared to the control (P < 0.001), indicating protection of the host against infection by A. hydrophila. In conclusion, secretory metabolites did not play major roles in the growth promotion and disease protection effects of yeast. Nevertheless, secretory metabolites were the major contributing factor towards improved gut microvilli morphology, relieved stress status, and reduced intestinal inflammation of Nile tilapia fed diets supplemented with baker’s yeast. PMID:26696403

Optimization of carbon and nitrogen medium components for biomass production using non-Saccharomyces wine yeasts.

PubMed

Schnierda, T; Bauer, F F; Divol, B; van Rensburg, E; Görgens, J F

2014-05-01

The impact of different nitrogen and carbon sources on biomass production of the non-Saccharomyces wine yeast species Lachancea thermotolerans, Metschnikowia pulcherrima and Issatchenkia orientalis was assessed. Using a molasses-based medium, yeast extract and corn steep liquor as well as ammonium sulphate and di-ammonium phosphate (DAP) as nitrogen sources were compared in shake-flask cultures. A medium with 20 g l⁻¹ sugar (diluted molasses) and 500 mg l⁻¹ total yeast assimilable nitrogen, from yeast extract, gave the highest biomass concentrations and yields. Invertase pretreatment was required for cultures of M. pulcherrima and I. orientalis, and respective biomass yields of 0.7 and 0.8 g g⁻¹ were achieved in aerobic bioreactor cultures. The absence of ethanol production suggested Crabtree-negative behaviour by these yeasts, whereas Crabtree-positive behaviour by L. thermotolerans resulted in ethanol and biomass concentrations of 5.5 and 11.1 g l⁻¹, respectively. Recent studies demonstrate that non-Saccharomyces yeasts confer positive attributes to the final composition of wine. However, optimal process conditions for their biomass production have not been described, thereby limiting commercial application. In this study, industrial media and methods of yeast cultivation were investigated to develop protocols for biomass production of non-Saccharomyces yeast starter cultures for the wine industry. © 2014 The Society for Applied Microbiology.

Establishment of intestinal microbiota with focus on yeasts of unweaned and weaned piglets kept under different farm conditions.

PubMed

Urubschurov, Vladimir; Janczyk, Pawel; Souffrant, Wolfgang-Bernhard; Freyer, Gertraude; Zeyner, Annette

2011-09-01

This study aimed to characterize the intestinal yeasts in weaning piglets and to establish their possible relationships with main bacterial groups. German Landrace piglets were weaned (WP, n=32) at 28 days of age or kept with the dams until day 39 without creep feed (UP, n=32). The experiment was performed at an experimental and a commercial farm (CF). Faeces were collected from the piglets, sows and pen floors on days 28, 33 and 39 for isolation of DNA and cultivation for enumeration of yeasts, enterobacteria, enterococci and lactobacilli. Fragments of the D1 domain of 26S rRNA gene were amplified and separated by denaturing gradient gel electrophoresis (DGGE). No yeasts could be cultured from water and feed samples. No or only low numbers of yeasts were detected among all UP. In WP at CF, yeasts correlated with lactobacilli (r=0.456; P=0.009) and enterobacteria (r=-0.407; P=0.021). Kazachstania slooffiae dominated among the cultured yeasts. It was the only yeast species detected by PCR-DGGE. Yeasts, especially K. slooffiae, established in the porcine gastrointestinal tract after consumption of grain-based feed and may interrelate with the intestinal microbiota. The study provides data indicating importance of K. slooffiae for the development of balanced porcine gut microbiota. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

Beta-glucan-depleted, glycopeptide-rich extracts from Brewer's and Baker's yeast (Saccharomyces cerevisiae) lower interferon-gamma production by stimulated human blood cells in vitro.

PubMed

Williams, Roderick; Dias, Daniel A; Jayasinghe, Nirupama; Roessner, Ute; Bennett, Louise E

2016-04-15

Regulation of the human immune system requires controlled pro- and anti-inflammatory responses for host defence against infection and disease states. Yeasts (Saccharomyces cerevisiae), as used in brewing and baking, are mostly known for ability to stimulate the human immune-system predominantly reflecting the pro-inflammatory cell wall β-glucans. However, in this study, using food-compatible processing methods, glycopeptide-enriched and β-glucan-depleted products were each prepared from Brewer's and Baker's yeasts, which suppressed production of interferon-γ (IFN-γ) in human whole blood cell assay, signifying that anti-inflammatory factors are also present in yeast. Anti-inflammatory bioactivities of products prepared from Brewer's and Baker's yeast were compared with the commercial yeast product, Epicor®. While unfractionated Epicor was inactive, the C18 resin-binding fractions of Brewer's and Baker's yeast products and Epicor dose-dependently lowered IFN-γ, demonstrating that Epicor also contained both pro-inflammatory (β-glucans) and anti-inflammatory components. Anti-inflammatory activity was attributed to C18 resin-binding species glyco-peptides in Epicor and experimental yeast products. This study demonstrated that pro- and anti-inflammatory factors could be resolved and enriched in yeasts by suitable processing, with potential to improve specific activities. Crown Copyright © 2015. Published by Elsevier Ltd. All rights reserved.

Study on the effects of near-future ocean acidification on marine yeasts: a microcosm approach

NASA Astrophysics Data System (ADS)

Krause, Evamaria; Wichels, Antje; Erler, René; Gerdts, Gunnar

2013-12-01

Marine yeasts play an important role in biodegradation and nutrient cycling and are often associated with marine flora and fauna. They show maximum growth at pH levels lower than present-day seawater pH. Thus, contrary to many other marine organisms, they may actually profit from ocean acidification. Hence, we conducted a microcosm study, incubating natural seawater from the North Sea at present-day pH (8.10) and two near-future pH levels (7.81 and 7.67). Yeasts were isolated from the initial seawater sample and after 2 and 4 weeks of incubation. Isolates were classified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and representative isolates were identified by partial sequencing of the large subunit rRNA gene. From the initial seawater sample, we predominantly isolated a yeast-like filamentous fungus related to Aureobasidium pullulans, Cryptococcus sp., Candida sake, and various cold-adapted yeasts. After incubation, we found more different yeast species at near-future pH levels than at present-day pH. Yeasts reacting to low pH were related to Leucosporidium scottii, Rhodotorula mucilaginosa, Cryptococcus sp., and Debaryomyces hansenii. Our results suggest that these yeasts will benefit from seawater pH reductions and give a first indication that the importance of yeasts will increase in a more acidic ocean.

Interactions between Drosophila and its natural yeast symbionts—Is Saccharomyces cerevisiae a good model for studying the fly-yeast relationship?

PubMed Central

Hoang, Don; Kopp, Artyom

2015-01-01

Yeasts play an important role in the biology of the fruit fly, Drosophila melanogaster. In addition to being a valuable source of nutrition, yeasts affect D. melanogaster behavior and interact with the host immune system. Most experiments investigating the role of yeasts in D. melanogaster biology use the baker’s yeast, Saccharomyces cerevisiae. However, S. cerevisiae is rarely found with natural populations of D. melanogaster or other Drosophila species. Moreover, the strain of S. cerevisiae used most often in D. melanogaster experiments is a commercially and industrially important strain that, to the best of our knowledge, was not isolated from flies. Since disrupting natural host–microbe interactions can have profound effects on host biology, the results from D. melanogaster–S. cerevisiae laboratory experiments may not be fully representative of host–microbe interactions in nature. In this study, we explore the D. melanogaster-yeast relationship using five different strains of yeast that were isolated from wild Drosophila populations. Ingested live yeasts have variable persistence in the D. melanogaster gastrointestinal tract. For example, Hanseniaspora occidentalis persists relative to S. cerevisiae, while Brettanomyces naardenensis is removed. Despite these differences in persistence relative to S. cerevisiae, we find that all yeasts decrease in total abundance over time. Reactive oxygen species (ROS) are an important component of the D. melanogaster anti-microbial response and can inhibit S. cerevisiae growth in the intestine. To determine if sensitivity to ROS explains the differences in yeast persistence, we measured yeast growth in the presence and absence of hydrogen peroxide. We find that B. naardenesis is completely inhibited by hydrogen peroxide, while H. occidentalis is not, which is consistent with yeast sensitivity to ROS affecting persistence within the D. melanogaster gastrointestinal tract. We also compared the feeding preference of D. melanogaster when given the choice between a naturally associated yeast and S. cerevisiae. We do not find a correlation between preferred yeasts and those that persist in the intestine. Notably, in no instances is S. cerevisiae preferred over the naturally associated strains. Overall, our results show that D. melanogaster-yeast interactions are more complex than might be revealed in experiments that use only S. cerevisiae. We propose that future research utilize other yeasts, and especially those that are naturally associated with Drosophila, to more fully understand the role of yeasts in Drosophila biology. Since the genetic basis of host–microbe interactions is shared across taxa and since many of these genes are initially discovered in D. melanogaster, a more realistic fly-yeast model system will benefit our understanding of host–microbe interactions throughout the animal kingdom. PMID:26336636

Interactions between Drosophila and its natural yeast symbionts-Is Saccharomyces cerevisiae a good model for studying the fly-yeast relationship?

PubMed

Hoang, Don; Kopp, Artyom; Chandler, James Angus

2015-01-01

Yeasts play an important role in the biology of the fruit fly, Drosophila melanogaster. In addition to being a valuable source of nutrition, yeasts affect D. melanogaster behavior and interact with the host immune system. Most experiments investigating the role of yeasts in D. melanogaster biology use the baker's yeast, Saccharomyces cerevisiae. However, S. cerevisiae is rarely found with natural populations of D. melanogaster or other Drosophila species. Moreover, the strain of S. cerevisiae used most often in D. melanogaster experiments is a commercially and industrially important strain that, to the best of our knowledge, was not isolated from flies. Since disrupting natural host-microbe interactions can have profound effects on host biology, the results from D. melanogaster-S. cerevisiae laboratory experiments may not be fully representative of host-microbe interactions in nature. In this study, we explore the D. melanogaster-yeast relationship using five different strains of yeast that were isolated from wild Drosophila populations. Ingested live yeasts have variable persistence in the D. melanogaster gastrointestinal tract. For example, Hanseniaspora occidentalis persists relative to S. cerevisiae, while Brettanomyces naardenensis is removed. Despite these differences in persistence relative to S. cerevisiae, we find that all yeasts decrease in total abundance over time. Reactive oxygen species (ROS) are an important component of the D. melanogaster anti-microbial response and can inhibit S. cerevisiae growth in the intestine. To determine if sensitivity to ROS explains the differences in yeast persistence, we measured yeast growth in the presence and absence of hydrogen peroxide. We find that B. naardenesis is completely inhibited by hydrogen peroxide, while H. occidentalis is not, which is consistent with yeast sensitivity to ROS affecting persistence within the D. melanogaster gastrointestinal tract. We also compared the feeding preference of D. melanogaster when given the choice between a naturally associated yeast and S. cerevisiae. We do not find a correlation between preferred yeasts and those that persist in the intestine. Notably, in no instances is S. cerevisiae preferred over the naturally associated strains. Overall, our results show that D. melanogaster-yeast interactions are more complex than might be revealed in experiments that use only S. cerevisiae. We propose that future research utilize other yeasts, and especially those that are naturally associated with Drosophila, to more fully understand the role of yeasts in Drosophila biology. Since the genetic basis of host-microbe interactions is shared across taxa and since many of these genes are initially discovered in D. melanogaster, a more realistic fly-yeast model system will benefit our understanding of host-microbe interactions throughout the animal kingdom.

Shock-wave equation-of-state measurements in fused silica up to 1600 GPa

DOE PAGES

McCoy, C. A.; Gregor, M. C.; Polsin, D. N.; ...

2016-06-02

The properties of silica are important to geophysical and high-pressure equation of state research. The most prevalent crystalline form, α-quartz, has been extensively studied to TPa pressures. Recent experiments with amorphous silica, commonly referred to as fused silica, provided Hugoniot and reflectivity data up to 630 GPa using magnetically-driven aluminum impactors. This article presents measurements of the fused silica Hugoniot over the range from 200 to 1600 GPa using laser-driven shocks with a quartz standard. These results extend the measured Hugoniot of fused silica to higher pressures, but more importantly, in the 200-600 GPa range, the data are very goodmore » agreement with those obtained with a different driver and standard material. As a result, a new shock velocity-particle velocity relation is derived to fit the experimental data.« less

Fused silica windows for solar receiver applications

NASA Astrophysics Data System (ADS)

Hertel, Johannes; Uhlig, Ralf; Söhn, Matthias; Schenk, Christian; Helsch, Gundula; Bornhöft, Hansjörg

2016-05-01

A comprehensive study of optical and mechanical properties of quartz glass (fused silica) with regard to application in high temperature solar receivers is presented. The dependence of rupture strength on different surface conditions as well as high temperature is analyzed, focussing particularly on damage by devitrification and sandblasting. The influence of typical types of contamination in combination with thermal cycling on the optical properties of fused silica is determined. Cleaning methods are compared regarding effectiveness on contamination-induced degradation for samples with and without antireflective coating. The FEM-aided design of different types of receiver windows and their support structure is presented. A large-scale production process has been developed for producing fused silica dome shaped windows (pressurized window) up to a diameter of 816 mm. Prototypes were successfully pressure-tested in a test bench and certified according to the European Pressure Vessel Directive.

Correlating yeast cell stress physiology to changes in the cell surface morphology: atomic force microscopic studies.

PubMed

Canetta, Elisabetta; Walker, Graeme M; Adya, Ashok K

2006-07-06

Atomic Force Microscopy (AFM) has emerged as a powerful biophysical tool in biotechnology and medicine to investigate the morphological, physical, and mechanical properties of yeasts and other biological systems. However, properties such as, yeasts' response to environmental stresses, metabolic activities of pathogenic yeasts, cell-cell/cell-substrate adhesion, and cell-flocculation have rarely been investigated so far by using biophysical tools. Our recent results obtained by AFM on one strain each of Saccharomyces cerevisiae and Schizosaccharomyces pombe show a clear correlation between the physiology of environmentally stressed yeasts and the changes in their surface morphology. The future directions of the AFM related techniques in relation to yeasts are also discussed.

The ecology and evolution of non-domesticated Saccharomyces species.

PubMed

Boynton, Primrose J; Greig, Duncan

2014-12-01

Yeast researchers need model systems for ecology and evolution, but the model yeast Saccharomyces cerevisiae is not ideal because its evolution has been affected by domestication. Instead, ecologists and evolutionary biologists are focusing on close relatives of S. cerevisiae, the seven species in the genus Saccharomyces. The best-studied Saccharomyces yeast, after S. cerevisiae, is S. paradoxus, an oak tree resident throughout the northern hemisphere. In addition, several more members of the genus Saccharomyces have recently been discovered. Some Saccharomyces species are only found in nature, while others include both wild and domesticated strains. Comparisons between domesticated and wild yeasts have pinpointed hybridization, introgression and high phenotypic diversity as signatures of domestication. But studies of wild Saccharomyces natural history, biogeography and ecology are only beginning. Much remains to be understood about wild yeasts' ecological interactions and life cycles in nature. We encourage researchers to continue to investigate Saccharomyces yeasts in nature, both to place S. cerevisiae biology into its ecological context and to develop the genus Saccharomyces as a model clade for ecology and evolution. © 2014 The Authors. Yeast published by John Wiley & Sons, Ltd.

The Yeast Saccharomyces cerevisiae as a Model for Understanding RAS Proteins and Their Role in Human Tumorigenesis

PubMed Central

Cazzanelli, Giulia; Francisco, Rita; Azevedo, Luísa; Carvalho, Patrícia Dias; Almeida, Ana; Côrte-Real, Manuela; Oliveira, Maria José; Lucas, Cândida; Sousa, Maria João

2018-01-01

The exploitation of the yeast Saccharomyces cerevisiae as a biological model for the investigation of complex molecular processes conserved in multicellular organisms, such as humans, has allowed fundamental biological discoveries. When comparing yeast and human proteins, it is clear that both amino acid sequences and protein functions are often very well conserved. One example of the high degree of conservation between human and yeast proteins is highlighted by the members of the RAS family. Indeed, the study of the signaling pathways regulated by RAS in yeast cells led to the discovery of properties that were often found interchangeable with RAS proto-oncogenes in human pathways, and vice versa. In this work, we performed an updated critical literature review on human and yeast RAS pathways, specifically highlighting the similarities and differences between them. Moreover, we emphasized the contribution of studying yeast RAS pathways for the understanding of human RAS and how this model organism can contribute to unveil the roles of RAS oncoproteins in the regulation of mechanisms important in the tumorigenic process, like autophagy. PMID:29463063

The ecology and evolution of non-domesticated Saccharomyces species

PubMed Central

Boynton, Primrose J; Greig, Duncan

2014-01-01

Yeast researchers need model systems for ecology and evolution, but the model yeast Saccharomyces cerevisiae is not ideal because its evolution has been affected by domestication. Instead, ecologists and evolutionary biologists are focusing on close relatives of S. cerevisiae, the seven species in the genus Saccharomyces. The best-studied Saccharomyces yeast, after S. cerevisiae, is S. paradoxus, an oak tree resident throughout the northern hemisphere. In addition, several more members of the genus Saccharomyces have recently been discovered. Some Saccharomyces species are only found in nature, while others include both wild and domesticated strains. Comparisons between domesticated and wild yeasts have pinpointed hybridization, introgression and high phenotypic diversity as signatures of domestication. But studies of wild Saccharomyces natural history, biogeography and ecology are only beginning. Much remains to be understood about wild yeasts' ecological interactions and life cycles in nature. We encourage researchers to continue to investigate Saccharomyces yeasts in nature, both to place S. cerevisiae biology into its ecological context and to develop the genus Saccharomyces as a model clade for ecology and evolution. © 2014 The Authors. Yeast published by John Wiley & Sons Ltd. PMID:25242436

Metabolomics-based prediction models of yeast strains for screening of metabolites contributing to ethanol stress tolerance

NASA Astrophysics Data System (ADS)

Hashim, Z.; Fukusaki, E.

2016-06-01

The increased demand for clean, sustainable and renewable energy resources has driven the development of various microbial systems to produce biofuels. One of such systems is the ethanol-producing yeast. Although yeast produces ethanol naturally using its native pathways, production yield is low and requires improvement for commercial biofuel production. Moreover, ethanol is toxic to yeast and thus ethanol tolerance should be improved to further enhance ethanol production. In this study, we employed metabolomics-based strategy using 30 single-gene deleted yeast strains to construct multivariate models for ethanol tolerance and screen metabolites that relate to ethanol sensitivity/tolerance. The information obtained from this study can be used as an input for strain improvement via metabolic engineering.

The hydrolytic activity of esterases in the yeast Saccharomyces cerevisiae is strain dependent.

PubMed

Kwolek-Mirek, Magdalena; Bednarska, Sabina; Zadrąg-Tęcza, Renata; Bartosz, Grzegorz

2011-11-01

Ester precursors of fluorogenic or chromogenic probes are often employed in studies of yeast cell biology. This study was aimed at a comparison of the ability of several commonly used laboratory wild-type Saccharomyces cerevisiae strains to hydrolyse the following model esters: fluorescein diacetate, 2-naphthyl acetate, PNPA (p-nitrophenyl acetate) and AMQI (7-acetoxy-1-methylquinolinum iodide). In all the strains, the esterase activity was localized mainly to the cytosol. Considerable differences in esterase activity were observed between various wild-type laboratory yeast strains. The phase of growth also contributed to the variation in esterase activity of the yeast. This diversity implies the need for caution in using intracellularly hydrolysed probes for a comparison of yeast strains with various genetic backgrounds.

End-joining inhibition at telomeres requires the translocase and polySUMO-dependent ubiquitin ligase Uls1.

PubMed

Lescasse, Rachel; Pobiega, Sabrina; Callebaut, Isabelle; Marcand, Stéphane

2013-03-20

In eukaryotes, permanent inhibition of the non-homologous end joining (NHEJ) repair pathway at telomeres ensures that chromosome ends do not fuse. In budding yeast, binding of Rap1 to telomere repeats establishes NHEJ inhibition. Here, we show that the Uls1 protein is required for the maintenance of NHEJ inhibition at telomeres. Uls1 protein is a non-essential Swi2/Snf2-related translocase and a Small Ubiquitin-related Modifier (SUMO)-Targeted Ubiquitin Ligase (STUbL) with unknown targets. Loss of Uls1 results in telomere-telomere fusions. Uls1 requirement is alleviated by the absence of poly-SUMO chains and by rap1 alleles lacking SUMOylation sites. Furthermore, Uls1 limits the accumulation of Rap1 poly-SUMO conjugates. We propose that one of Uls1 functions is to clear non-functional poly-SUMOylated Rap1 molecules from telomeres to ensure the continuous efficiency of NHEJ inhibition. Since Uls1 is the only known STUbL with a translocase activity, it can be the general molecular sweeper for the clearance of poly-SUMOylated proteins on DNA in eukaryotes.

Schizosaccharomyces pombe Retrotransposon Tf2 Mobilizes Primarily through Homologous cDNA Recombination

PubMed Central

Hoff, Eleanor F.; Levin, Henry L.; Boeke, Jef D.

1998-01-01

The Tf2 retrotransposon, found in the fission yeast Schizosaccharomyces pombe, is nearly identical to its sister element, Tf1, in its reverse transcriptase-RNase H and integrase domains but is very divergent in the gag domain, the protease, the 5′ untranslated region, and the U3 domain of the long terminal repeats. It has now been demonstrated that a neo-marked copy of Tf2 overexpressed from a heterologous promoter can mobilize into the S. pombe genome and produce true transposition events. However, the Tf2-neo mobilization frequency is 10- to 20-fold lower than that of Tf1-neo, and 70% of the Tf2-neo events are homologous recombination events generated independently of a functional Tf2 integrase. Thus, the Tf2 element is primarily dependent on homologous recombination with preexisting copies of Tf2 for its propagation. Finally, production of Tf2-neo proteins and cDNA was also analyzed; surprisingly, Tf2 was found to produce its reverse transcriptase as a single species in which it is fused to protease, unlike all other retroviruses and retrotransposons. PMID:9774697

Pooled-matrix protein interaction screens using Barcode Fusion Genetics.

PubMed

Yachie, Nozomu; Petsalaki, Evangelia; Mellor, Joseph C; Weile, Jochen; Jacob, Yves; Verby, Marta; Ozturk, Sedide B; Li, Siyang; Cote, Atina G; Mosca, Roberto; Knapp, Jennifer J; Ko, Minjeong; Yu, Analyn; Gebbia, Marinella; Sahni, Nidhi; Yi, Song; Tyagi, Tanya; Sheykhkarimli, Dayag; Roth, Jonathan F; Wong, Cassandra; Musa, Louai; Snider, Jamie; Liu, Yi-Chun; Yu, Haiyuan; Braun, Pascal; Stagljar, Igor; Hao, Tong; Calderwood, Michael A; Pelletier, Laurence; Aloy, Patrick; Hill, David E; Vidal, Marc; Roth, Frederick P

2016-04-22

High-throughput binary protein interaction mapping is continuing to extend our understanding of cellular function and disease mechanisms. However, we remain one or two orders of magnitude away from a complete interaction map for humans and other major model organisms. Completion will require screening at substantially larger scales with many complementary assays, requiring further efficiency gains in proteome-scale interaction mapping. Here, we report Barcode Fusion Genetics-Yeast Two-Hybrid (BFG-Y2H), by which a full matrix of protein pairs can be screened in a single multiplexed strain pool. BFG-Y2H uses Cre recombination to fuse DNA barcodes from distinct plasmids, generating chimeric protein-pair barcodes that can be quantified via next-generation sequencing. We applied BFG-Y2H to four different matrices ranging in scale from ~25 K to 2.5 M protein pairs. The results show that BFG-Y2H increases the efficiency of protein matrix screening, with quality that is on par with state-of-the-art Y2H methods. © 2016 The Authors. Published under the terms of the CC BY 4.0 license.

Inner hydrogen atom transfer in benzo-fused low symmetrical metal-free tetraazaporphyrin and phthalocyanine analogues: density functional theory studies.

PubMed

Qi, Dongdong; Zhang, Yuexing; Cai, Xue; Jiang, Jianzhuang; Bai, Ming

2009-02-01

Density functional theory (DFT) calculations were carried out to study the inner hydrogen atom transfer in low symmetrical metal-free tetrapyrrole analogues ranging from tetraazaporphyrin H(2)TAP (A(0)B(0)C(0)D(0)) to naphthalocyanine H(2)Nc (A(2)B(2)C(2)D(2)) via phthalocyanine H(2)Pc (A(1)B(1)C(1)D(1)). All the transition paths of sixteen different compounds (A(0)B(0)C(0)D(0)-A(2)B(2)C(2)D(2) and A(0)B(0)C(m)D(n), m

Meta-analysis of the influence of Saccharomyces cerevisiae supplementation on ruminal parameters and milk production of ruminants.

PubMed

Desnoyers, M; Giger-Reverdin, S; Bertin, G; Duvaux-Ponter, C; Sauvant, D

2009-04-01

The effects of yeast supplementation on intake, production, and rumen fermentation characteristics have been widely studied, but results are inconsistent between different studies. A quantitative meta-analysis was applied to 110 papers, 157 experiments, and 376 treatments dealing with yeast supplementation in ruminants. The objective was first to highlight the major quantitative effects of live yeast supplementation on intake, rumen fermentation, and milk production, and second, to identify major differences in experimental conditions between studies that can affect the response to treatment. Some of these experimental conditions are referred to as interfering factors. Yeast supplementation increased rumen pH (+0.03 on average) and rumen volatile fatty acid concentration (+2.17 mM on average), tended to decrease rumen lactic acid concentration (-0.9 mM on average), and had no influence on acetate-to-propionate ratio. Total-tract organic matter digestibility was also increased by yeast supplementation (+0.8% on average). Yeast supplementation increased dry matter intake (DMI; +0.44 g/kg of body weight; BW), milk yield (+1.2 g/kg of BW), and tended to increase milk fat content (+0.05%), but had no influence on milk protein content. Dose effects of yeast supplementation, expressed as log(10) [1+(cfu per 100 kg of BW)], globally confirmed the qualitative effects observed in the first analysis. The positive effect of yeast supplementation on rumen pH increased with the percentage of concentrate in the diet and with the DMI level. It was negatively correlated with the level of dietary neutral detergent fiber (NDF). The positive effect of yeast supplementation on rumen volatile fatty acid concentration increased with DMI and crude protein levels. The positive effect of yeast supplementation on organic matter digestibility increased with the percentage of concentrate and NDF in the diet. The negative effect of yeast supplementation on lactic acid concentration tended to decrease when the DMI level and the percentage of concentrate in the diet increased. The effects of interfering factors were globally similar when either dose effect or qualitative effect of yeast was taken into account. Although rumen fermentation efficiency per se was not measured, these results suggest an improvement in rumen fermentation by yeast supplementation. This effect could, however, be modulated by several different factors such as DMI, percentage of concentrate or NDF in the diet, or species.

Transformation of Mycelial and Yeast Forms of Paracoccidioides brasiliensis in Cultures and in Experimental Inoculations

PubMed Central

Carbonell, Luis M.; Rodríguez, Joaquín

1965-01-01

Carbonell, Luis M. (Instituto Venezolano de Investigaciones Científicas, Caracas, Venezuela), and Joaquín Rodríguez. Transformation of mycelial and yeast forms of Paracoccidioides brasiliensis in cultures and in experimental inoculations. J. Bacteriol. 90:504–510. 1965.—Experimental transformations of mycelial to yeast and yeast to mycelial forms in culture, and mycelial to yeast forms in tissue, were studied. All the transitional forms that appeared in culture were also seen in tissue, but in fewer number. Most of the hyphae in culture were transformed into yeast, but only a few in tissue. Yeast appeared in testicle around the 3rd day after inoculation, but on the 10th day in subcutaneous tissue. Pathogenicity of mycelium was high, since yeast was found in almost all of the organs inoculated with mycelium. Histologically, an acute inflammation occurred first, owing to the inoculation of mycelium, followed by a giant-cell granuloma with abundant hyphae detritus. These giant cells almost disappeared about 10 days after inoculation, giving place to a second giant-cell granuloma with yeast forms. Images PMID:14329466

Analysis of non-Saccharomyces yeast populations isolated from grape musts from Sicily (Italy).

PubMed

Romancino, D P; Di Maio, S; Muriella, R; Oliva, D

2008-12-01

The aim of this study was to identify the non-Saccharomyces yeast populations present in the grape must microflora from wineries from different areas around the island of Sicily. Yeasts identification was conducted on 2575 colonies isolated from six musts, characterized using Wallerstein Laboratory (WL) nutrient agar, restriction analysis of the amplified 5.8S-internal transcribed spacer region and restriction profiles of amplified 26S rDNA. In those colonies, we identified 11 different yeast species originating from wine musts from two different geographical areas of the island of Sicily. We isolated non-Saccharomyces yeasts and described the microflora in grape musts from different areas of Sicily. Moreover, we discovered two new colony morphologies for yeasts on WL agar never previously described. This investigation is a first step in understanding the distribution of non-Saccharomyces yeasts in grape musts from Sicily. The contribution is important as a tool for monitoring the microflora in grape musts and for establishing a new non-Saccharomyces yeast collection; in the future, this collection will be used for understanding the significance of these yeasts in oenology.

A polyphasic study on the taxonomic position of industrial sour dough yeasts.

PubMed

Mäntynen, V H; Korhola, M; Gudmundsson, H; Turakainen, H; Alfredsson, G A; Salovaara, H; Lindström, K

1999-02-01

The sour dough bread making process is extensively used to produce wholesome palatable rye bread. The process is traditionally done using a back-slopping procedure. Traditional sour doughs in Finland comprise of lactic acid bacteria and yeasts. The yeasts present in these doughs have been enriched in the doughs due to their metabolic activities, e.g. acid tolerance. We characterized the yeasts in five major sour bread bakeries in Finland. We found that most of the commercial sour doughs contained yeasts which were similar to Candida milleri on the basis of 18S rDNA and EF-3 PCR-RFLP patterns and metabolic activities. Some of the bakery yeasts exhibited extensive karyotype polymorphism. The minimum growth temperature was 8 degrees C for C. milleri and also for most of sour dough yeasts.

Metabolic Engineering of Oleaginous Yeasts for Production of Fuels and Chemicals

PubMed Central

Shi, Shuobo; Zhao, Huimin

2017-01-01

Oleaginous yeasts have been increasingly explored for production of chemicals and fuels via metabolic engineering. Particularly, there is a growing interest in using oleaginous yeasts for the synthesis of lipid-related products due to their high lipogenesis capability, robustness, and ability to utilize a variety of substrates. Most of the metabolic engineering studies in oleaginous yeasts focused on Yarrowia that already has plenty of genetic engineering tools. However, recent advances in systems biology and synthetic biology have provided new strategies and tools to engineer those oleaginous yeasts that have naturally high lipid accumulation but lack genetic tools, such as Rhodosporidium, Trichosporon, and Lipomyces. This review highlights recent accomplishments in metabolic engineering of oleaginous yeasts and recent advances in the development of genetic engineering tools in oleaginous yeasts within the last 3 years. PMID:29167664

Phenotypic landscape of non-conventional yeast species for different stress tolerance traits desirable in bioethanol fermentation.

PubMed

Mukherjee, Vaskar; Radecka, Dorota; Aerts, Guido; Verstrepen, Kevin J; Lievens, Bart; Thevelein, Johan M

2017-01-01

Non-conventional yeasts present a huge, yet barely exploited, resource of yeast biodiversity for industrial applications. This presents a great opportunity to explore alternative ethanol-fermenting yeasts that are more adapted to some of the stress factors present in the harsh environmental conditions in second-generation (2G) bioethanol fermentation. Extremely tolerant yeast species are interesting candidates to investigate the underlying tolerance mechanisms and to identify genes that when transferred to existing industrial strains could help to design more stress-tolerant cell factories. For this purpose, we performed a high-throughput phenotypic evaluation of a large collection of non-conventional yeast species to identify the tolerance limits of the different yeast species for desirable stress tolerance traits in 2G bioethanol production. Next, 12 multi-tolerant strains were selected and used in fermentations under different stressful conditions. Five strains out of which, showing desirable fermentation characteristics, were then evaluated in small-scale, semi-anaerobic fermentations with lignocellulose hydrolysates. Our results revealed the phenotypic landscape of many non-conventional yeast species which have not been previously characterized for tolerance to stress conditions relevant for bioethanol production. This has identified for each stress condition evaluated several extremely tolerant non- Saccharomyces yeasts. It also revealed multi-tolerance in several yeast species, which makes those species good candidates to investigate the molecular basis of a robust general stress tolerance. The results showed that some non-conventional yeast species have similar or even better fermentation efficiency compared to S. cerevisiae in the presence of certain stressful conditions. Prior to this study, our knowledge on extreme stress-tolerant phenotypes in non-conventional yeasts was limited to only few species. Our work has now revealed in a systematic way the potential of non- Saccharomyces species to emerge either as alternative host species or as a source of valuable genetic information for construction of more robust industrial S. serevisiae bioethanol production yeasts. Striking examples include yeast species like Pichia kudriavzevii and Wickerhamomyces anomalus that show very high tolerance to diverse stress factors. This large-scale phenotypic analysis has yielded a detailed database useful as a resource for future studies to understand and benefit from the molecular mechanisms underlying the extreme phenotypes of non-conventional yeast species.

Non-Saccharomyces Yeasts Nitrogen Source Preferences: Impact on Sequential Fermentation and Wine Volatile Compounds Profile

PubMed Central

Gobert, Antoine; Tourdot-Maréchal, Raphaëlle; Morge, Christophe; Sparrow, Céline; Liu, Youzhong; Quintanilla-Casas, Beatriz; Vichi, Stefania; Alexandre, Hervé

2017-01-01

Nitrogen sources in the must are important for yeast metabolism, growth, and performance, and wine volatile compounds profile. Yeast assimilable nitrogen (YAN) deficiencies in grape must are one of the main causes of stuck and sluggish fermentation. The nitrogen requirement of Saccharomyces cerevisiae metabolism has been described in detail. However, the YAN preferences of non-Saccharomyces yeasts remain unknown despite their increasingly widespread use in winemaking. Furthermore, the impact of nitrogen consumption by non-Saccharomyces yeasts on YAN availability, alcoholic performance and volatile compounds production by S. cerevisiae in sequential fermentation has been little studied. With a view to improving the use of non-Saccharomyces yeasts in winemaking, we studied the use of amino acids and ammonium by three strains of non-Saccharomyces yeasts (Starmerella bacillaris, Metschnikowia pulcherrima, and Pichia membranifaciens) in grape juice. We first determined which nitrogen sources were preferentially used by these yeasts in pure cultures at 28 and 20°C (because few data are available). We then carried out sequential fermentations at 20°C with S. cerevisiae, to assess the impact of the non-Saccharomyces yeasts on the availability of assimilable nitrogen for S. cerevisiae. Finally, 22 volatile compounds were quantified in sequential fermentation and their levels compared with those in pure cultures of S. cerevisiae. We report here, for the first time, that non-Saccharomyces yeasts have specific amino-acid consumption profiles. Histidine, methionine, threonine, and tyrosine were not consumed by S. bacillaris, aspartic acid was assimilated very slowly by M. pulcherrima, and glutamine was not assimilated by P. membranifaciens. By contrast, cysteine appeared to be a preferred nitrogen source for all non-Saccharomyces yeasts. In sequential fermentation, these specific profiles of amino-acid consumption by non-Saccharomyces yeasts may account for some of the interactions observed here, such as poorer performances of S. cerevisiae and volatile profile changes. PMID:29163451

Non-Saccharomyces Yeasts Nitrogen Source Preferences: Impact on Sequential Fermentation and Wine Volatile Compounds Profile.

PubMed

Gobert, Antoine; Tourdot-Maréchal, Raphaëlle; Morge, Christophe; Sparrow, Céline; Liu, Youzhong; Quintanilla-Casas, Beatriz; Vichi, Stefania; Alexandre, Hervé

2017-01-01

Nitrogen sources in the must are important for yeast metabolism, growth, and performance, and wine volatile compounds profile. Yeast assimilable nitrogen (YAN) deficiencies in grape must are one of the main causes of stuck and sluggish fermentation. The nitrogen requirement of Saccharomyces cerevisiae metabolism has been described in detail. However, the YAN preferences of non- Saccharomyces yeasts remain unknown despite their increasingly widespread use in winemaking. Furthermore, the impact of nitrogen consumption by non- Saccharomyces yeasts on YAN availability, alcoholic performance and volatile compounds production by S. cerevisiae in sequential fermentation has been little studied. With a view to improving the use of non- Saccharomyces yeasts in winemaking, we studied the use of amino acids and ammonium by three strains of non- Saccharomyces yeasts ( Starmerella bacillaris, Metschnikowia pulcherrima , and Pichia membranifaciens ) in grape juice. We first determined which nitrogen sources were preferentially used by these yeasts in pure cultures at 28 and 20°C (because few data are available). We then carried out sequential fermentations at 20°C with S. cerevisiae , to assess the impact of the non- Saccharomyces yeasts on the availability of assimilable nitrogen for S. cerevisiae . Finally, 22 volatile compounds were quantified in sequential fermentation and their levels compared with those in pure cultures of S. cerevisiae . We report here, for the first time, that non- Saccharomyces yeasts have specific amino-acid consumption profiles. Histidine, methionine, threonine, and tyrosine were not consumed by S. bacillaris , aspartic acid was assimilated very slowly by M. pulcherrima , and glutamine was not assimilated by P. membranifaciens . By contrast, cysteine appeared to be a preferred nitrogen source for all non- Saccharomyces yeasts. In sequential fermentation, these specific profiles of amino-acid consumption by non- Saccharomyces yeasts may account for some of the interactions observed here, such as poorer performances of S. cerevisiae and volatile profile changes.

Chromosomal Aneuploidy Improves the Brewing Characteristics of Sake Yeast.

PubMed

Kadowaki, Masafumi; Fujimaru, Yuki; Taguchi, Seiga; Ferdouse, Jannatul; Sawada, Kazutaka; Kimura, Yuta; Terasawa, Yohei; Agrimi, Gennaro; Anai, Toyoaki; Noguchi, Hideki; Toyoda, Atsushi; Fujiyama, Asao; Akao, Takeshi; Kitagaki, Hiroshi

2017-12-15

The effect of chromosomal aneuploidy on the brewing characteristics of brewery yeasts has not been studied. Here we report that chromosomal aneuploidy in sake brewery yeast ( Saccharomyces cerevisiae ) leads to the development of favorable brewing characteristics. We found that pyruvate-underproducing sake yeast, which produces less off-flavor diacetyl, is aneuploid and trisomic for chromosomes XI and XIV. To confirm that this phenotype is due to aneuploidy, we obtained 45 haploids with various chromosomal additions and investigated their brewing profiles. A greater number of chromosomes correlated with a decrease in pyruvate production. Especially, sake yeast haploids with extra chromosomes in addition to chromosome XI produced less pyruvate than euploids. Mitochondrion-related metabolites and intracellular oxygen species in chromosome XI aneuploids were higher than those in euploids, and this effect was canceled in their "petite" strains, suggesting that an increase in chromosomes upregulated mitochondrial activity and decreased pyruvate levels. These findings suggested that an increase in chromosome number, including chromosome XI, in sake yeast haploids leads to pyruvate underproduction through the augmentation of mitochondrial activity. This is the first report proposing that aneuploidy in brewery yeasts improves their brewing profile. IMPORTANCE Chromosomal aneuploidy has not been evaluated in development of sake brewing yeast strains. This study shows the relationship between chromosomal aneuploidy and brewing characteristics of brewery yeast strains. High concentrations of pyruvate during sake storage give rise to α-acetolactate and, in turn, to high concentrations of diacetyl, which is considered an off-flavor. It was demonstrated that pyruvate-underproducing sake yeast is trisomic for chromosome XI and XIV. Furthermore, sake yeast haploids with extra chromosomes produced reduced levels of pyruvate and showed metabolic processes characteristic of increased mitochondrial activity. This novel discovery will enable the selection of favorable brewery yeasts by monitoring the copy numbers of specific chromosomes through a process that does not involve generation/use of genetically modified organisms. Copyright © 2017 American Society for Microbiology.

Chromosomal Aneuploidy Improves the Brewing Characteristics of Sake Yeast

PubMed Central

Kadowaki, Masafumi; Fujimaru, Yuki; Taguchi, Seiga; Ferdouse, Jannatul; Sawada, Kazutaka; Kimura, Yuta; Terasawa, Yohei; Agrimi, Gennaro; Anai, Toyoaki; Noguchi, Hideki; Toyoda, Atsushi; Fujiyama, Asao; Akao, Takeshi

2017-01-01

ABSTRACT The effect of chromosomal aneuploidy on the brewing characteristics of brewery yeasts has not been studied. Here we report that chromosomal aneuploidy in sake brewery yeast (Saccharomyces cerevisiae) leads to the development of favorable brewing characteristics. We found that pyruvate-underproducing sake yeast, which produces less off-flavor diacetyl, is aneuploid and trisomic for chromosomes XI and XIV. To confirm that this phenotype is due to aneuploidy, we obtained 45 haploids with various chromosomal additions and investigated their brewing profiles. A greater number of chromosomes correlated with a decrease in pyruvate production. Especially, sake yeast haploids with extra chromosomes in addition to chromosome XI produced less pyruvate than euploids. Mitochondrion-related metabolites and intracellular oxygen species in chromosome XI aneuploids were higher than those in euploids, and this effect was canceled in their “petite” strains, suggesting that an increase in chromosomes upregulated mitochondrial activity and decreased pyruvate levels. These findings suggested that an increase in chromosome number, including chromosome XI, in sake yeast haploids leads to pyruvate underproduction through the augmentation of mitochondrial activity. This is the first report proposing that aneuploidy in brewery yeasts improves their brewing profile. IMPORTANCE Chromosomal aneuploidy has not been evaluated in development of sake brewing yeast strains. This study shows the relationship between chromosomal aneuploidy and brewing characteristics of brewery yeast strains. High concentrations of pyruvate during sake storage give rise to α-acetolactate and, in turn, to high concentrations of diacetyl, which is considered an off-flavor. It was demonstrated that pyruvate-underproducing sake yeast is trisomic for chromosome XI and XIV. Furthermore, sake yeast haploids with extra chromosomes produced reduced levels of pyruvate and showed metabolic processes characteristic of increased mitochondrial activity. This novel discovery will enable the selection of favorable brewery yeasts by monitoring the copy numbers of specific chromosomes through a process that does not involve generation/use of genetically modified organisms. PMID:28986374

KSC-99pp0702

NASA Image and Video Library

1999-06-17

At Launch Pad 17A, Cape Canaveral Air Station (CCAS), workers check out the protective cover placed over the top of NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite. The satellite is scheduled to be launched from CCAS June 23 aboard a Boeing Delta II rocket. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe

Fusion of arkosic sand by intrusive andesite

USGS Publications Warehouse

Bailey, Roy A.

1954-01-01

An andesite dike in the Valles Mountains of northern New Mexico has intruded and partly fused arkosic sediments for a distance of 50 feet from its contacts. The dike is semi-circular in form, has a maximum width of about 100 feet, and is about 500 feet long. Small associated arcuate dikes are arranged in spiral fashion around the main dike, suggesting that they were intruded along shear fractures similar to those described by Burbank (1941). The fused rocks surrounding the andesite dike are of three general types: 1) partly fused arkosic sand, 2) fused clay, and 3) hybrid rocks. The fused arkosic sand consists of relict detrital grains of quartz, orthoclose, and plagioclase, imbedded in colorless glass containing microlites of tridymite, cordierite, and magnetite. The relict quartz grains are corroded and embayed by glass; the orthoclase is sanidinized and partly fused; and the plagioclase is inverted to the high temperature form and is partly fused. The fused clay, which was originally a mixture of montmorillonite and hydromica, consists primarily of cordierite but also contains needle-like crystals of sillimanite (?) or mullite (?). The hybrid rocks originated in part by intermixing of fused arkosic sediments and andesitic liquid and in part by diffusion of mafic constituents through the fused sediments. They are rich in cordierite and magnetite and also contain hypersthene, augite, and plagioclase. The composition of pigeonite in the andesite indicates that the temperature of the andesite at the time of intrusion probably did not exceed 1200?C. Samples of arkosic sand were fused in the presence of water in a Morey bomb at 1050?C. Stability relations of certain minerals in the fused sand suggest that fusion may have taken place at a lower temperature, however, and the fluxing action of volatiles from the andesite are thought to have made this possible.

Pixel-based image fusion with false color mapping

NASA Astrophysics Data System (ADS)

Zhao, Wei; Mao, Shiyi

2003-06-01

In this paper, we propose a pixel-based image fusion algorithm that combines the gray-level image fusion method with the false color mapping. This algorithm integrates two gray-level images presenting different sensor modalities or at different frequencies and produces a fused false-color image. The resulting image has higher information content than each of the original images. The objects in the fused color image are easy to be recognized. This algorithm has three steps: first, obtaining the fused gray-level image of two original images; second, giving the generalized high-boost filtering images between fused gray-level image and two source images respectively; third, generating the fused false-color image. We use the hybrid averaging and selection fusion method to obtain the fused gray-level image. The fused gray-level image will provide better details than two original images and reduce noise at the same time. But the fused gray-level image can't contain all detail information in two source images. At the same time, the details in gray-level image cannot be discerned as easy as in a color image. So a color fused image is necessary. In order to create color variation and enhance details in the final fusion image, we produce three generalized high-boost filtering images. These three images are displayed through red, green and blue channel respectively. A fused color image is produced finally. This method is used to fuse two SAR images acquired on the San Francisco area (California, USA). The result shows that fused false-color image enhances the visibility of certain details. The resolution of the final false-color image is the same as the resolution of the input images.

Oleaginous yeasts for biodiesel: current and future trends in biology and production.

PubMed

Sitepu, Irnayuli R; Garay, Luis A; Sestric, Ryan; Levin, David; Block, David E; German, J Bruce; Boundy-Mills, Kyria L

2014-11-15

Production of biodiesel from edible plant oils is quickly expanding worldwide to fill a need for renewable, environmentally-friendly liquid transportation fuels. Due to concerns over use of edible commodities for fuels, production of biodiesel from non-edible oils including microbial oils is being developed. Microalgae biodiesel is approaching commercial viability, but has some inherent limitations such as requirements for sunlight. While yeast oils have been studied for decades, recent years have seen significant developments including discovery of new oleaginous yeast species and strains, greater understanding of the metabolic pathways that determine oleaginicity, optimization of cultivation processes for conversion of various types of waste plant biomass to oil using oleaginous yeasts, and development of strains with enhanced oil production. This review examines aspects of oleaginous yeasts not covered in depth in other recent reviews. Topics include the history of oleaginous yeast research, especially advances in the early 20th century; the phylogenetic diversity of oleaginous species, beyond the few species commonly studied; and physiological characteristics that should be considered when choosing yeast species and strains to be utilized for conversion of a given type of plant biomass to oleochemicals. Standardized terms are proposed for units that describe yeast cell mass and lipid production. Copyright © 2014. Published by Elsevier Inc.

Identification of oleaginous yeast strains able to accumulate high intracellular lipids when cultivated in alkaline pretreated corn stover

PubMed Central

Sitepu, Irnayuli R.; Jin, Mingjie; Fernandez, J. Enrique; da Costa Sousa, Leonardo; Balan, Venkatesh; Boundy-Mills, Kyria L.

2015-01-01

Microbial oil is a potential alternative to food/plant-derived biodiesel fuel. Our previous screening studies identified a wide range of oleaginous yeast species, using a defined laboratory medium known to stimulate lipid accumulation. In this study, the ability of these yeasts to grow and accumulate lipids was further investigated in synthetic hydrolysate (SynH) and authentic ammonia fiber expansion (AFEX™)-pretreated corn stover hydrolysate (ACSH). Most yeast strains tested were able to accumulate lipids in SynH, but only a few were able to grow and accumulate lipids in ACSH medium. Cryptococcus humicola UCDFST 10-1004 was able to accumulate as high as 15.5 g/L lipids, out of a total of 36 g/L cellular biomass when grown in ACSH, with a cellular lipid content of 40% of cell dry weight. This lipid production is among the highest reported values for oleaginous yeasts grown in authentic hydrolysate. Pre-culturing in SynH media with xylose as sole carbon source enabled yeasts to assimilate both glucose and xylose more efficiently in the subsequent hydrolysate medium. This study demonstrates that ACSH is a suitable medium for certain oleaginous yeasts to convert lignocellullosic sugars to triacylglycerols for production of biodiesel and other valuable oleochemicals. PMID:25052467

Oral yeast colonization throughout pregnancy.

PubMed

Rio, R; Simões-Silva, L; Garro, S; Silva, M-J; Azevedo, Á; Sampaio-Maia, B

2017-03-01

Recent studies suggest that placenta may harbour a unique microbiome that may have origin in maternal oral microbiome. Although the major physiological and hormonal adjustments observed in pregnant women lead to biochemical and microbiological modifications of the oral environment, very few studies evaluated the changes suffered by the oral microbiota throughout pregnancy. So, the aim of our study was to evaluate oral yeast colonization throughout pregnancy and to compare it with non-pregnant women. The oral yeast colonization was assessed in saliva of 30 pregnant and non-pregnant women longitudinally over a 6-months period. Demographic information was collected, a non-invasive intra-oral examination was performed and saliva flow and pH were determined. Pregnant and non-pregnant groups were similar regarding age and level of education. Saliva flow rate did not differ, but saliva pH was lower in pregnant than in non-pregnant women. Oral yeast prevalence was higher in pregnant than in non-pregnant women, either in the first or in the third trimester, but did not attain statistical significance. In individuals colonized with yeast, the total yeast quantification (Log10CFU/mL) increase from the 1st to the 3rd trimester in pregnant women, but not in non-pregnant women. Pregnancy may favour oral yeast growth that may be associated with an acidic oral environment.

Isolation, identification and growth determination of lactic acid-utilizing yeasts from the ruminal fluid of dairy cattle.

PubMed

Sirisan, V; Pattarajinda, V; Vichitphan, K; Leesing, R

2013-08-01

Ruminal organic acid production, especially lactic acid, can be modified by feeding cattle highly concentrated diets, which have been shown to adversely affect dairy cattle health. Therefore, the use of lactic acid-utilizing organisms is considered to be a potential method for controlling lactic acid levels. This study was conducted to isolate and identify lactic acid-utilizing yeasts from the ruminal fluid of dairy cattle and to determine the specific growth rate and generation time when using lactic acid as a carbon source instead of glucose. Seventeen yeast isolates were examined in this study. Yeasts isolated from dairy cattle that were fed a high cassava pulp diet (HCP) had higher specific growth rates and shorter generation times than yeasts isolated from dairy cattle that were fed a high-concentrate diet (HC) and a mixed diet (M). The three most effective yeasts in terms of specific growth rate and generation time were Pichia kudriavzevii, Candida rugosa and Kodamaea ohmeri, with 99, 100 and 99% nucleotide identities, respectively. These three isolates could be used as potential probiotics in dairy cattle diets. This study demonstrates that yeasts isolated from the ruminal fluid of dairy cattle can utilize lactic acid as a carbon and energy source for growth. The isolated yeasts can be used as probiotic supplements for dairy cattle that are fed highly concentrated diets to reduce ruminal lactic acid production. © 2013 The Society for Applied Microbiology.

Vitality Stains and Real Time PCR Studies to Delineate the Interactions of Pichia anomala and Aspergillus flavus

USDA-ARS?s Scientific Manuscript database

The objectives of this study were to probe the effect of the yeast, P. anomala against A flavus by using real time RT-PCR technique and vitality fluorescent stains. Yeast and fungi were inoculated into a 250 ml-flask containing 50 ml potato dextrose broth (PDB) at yeast to fungus (Y : F) ratios of ...

Budding yeast chromatin is dispersed in a crowded nucleoplasm in vivo

PubMed Central

Chen, Chen; Lim, Hong Hwa; Shi, Jian; Tamura, Sachiko; Maeshima, Kazuhiro; Surana, Uttam; Gan, Lu

2016-01-01

Chromatin organization has an important role in the regulation of eukaryotic systems. Although recent studies have refined the three-dimensional models of chromatin organization with high resolution at the genome sequence level, little is known about how the most fundamental units of chromatin—nucleosomes—are positioned in three dimensions in vivo. Here we use electron cryotomography to study chromatin organization in the budding yeast Saccharomyces cerevisiae. Direct visualization of yeast nuclear densities shows no evidence of 30-nm fibers. Aside from preribosomes and spindle microtubules, few nuclear structures are larger than a tetranucleosome. Yeast chromatin does not form compact structures in interphase or mitosis and is consistent with being in an “open” configuration that is conducive to high levels of transcription. From our study and those of others, we propose that yeast can regulate its transcription using local nucleosome–nucleosome associations. PMID:27605704

[Effect of Invasive Herb Species on the Structure of Soil Yeast Complexes in Mixed Forests Exemplified by Impatiens parviflora DC].

PubMed

Glushakova, A M; Kachalkin, A V; Chernov, I Yu

2015-01-01

Yeast abundance and diversity in a mixed forest sod-podzol soil under Impatiens parviflora DC plants was studied in comparison with unimpaired aboriginal herbaceous plants typical of the Mid-Russian secondary, after-forest meadow. The study was carried out throughout the vegetation period. Standard microbiological plating techniques revealed 36 yeast species. Typical pedobiotic (Cryptococcus podzolicus, Wickerhamomyces anomalus) and eurybiotic yeast species (Rhodotorula mucilaginosa) predominated in both biotopes. The relative abundance of the autochthonous soil yeast species Cryptococcus podzolicus was higher in the soil under aboriginal herbs than under Impatiens parviflora. Sites with aboriginal vegetation were also characterized by high abundance of the pedogamous species Schwanniomyces castelli and Torulaspora delbrueckii. The share of yeastlike Trichosporon fungi with high hydrolytic activity was considerably higher under adventitious plants Impatiens parviflora, as well as in the previously studied soil under Heracleum sosnowskyi.

Direct concentration and viability measurement of yeast in corn mash using a novel imaging cytometry method.

PubMed

Chan, Leo L; Lyettefi, Emily J; Pirani, Alnoor; Smith, Tim; Qiu, Jean; Lin, Bo

2011-08-01

Worldwide awareness of fossil-fuel depletion and global warming has been increasing over the last 30 years. Numerous countries, including the USA and Brazil, have introduced large-scale industrial fermentation facilities for bioethanol, biobutanol, or biodiesel production. Most of these biofuel facilities perform fermentation using standard baker's yeasts that ferment sugar present in corn mash, sugar cane, or other glucose media. In research and development in the biofuel industry, selection of yeast strains (for higher ethanol tolerance) and fermentation conditions (yeast concentration, temperature, pH, nutrients, etc.) can be studied to optimize fermentation performance. Yeast viability measurement is needed to identify higher ethanol-tolerant yeast strains, which may prolong the fermentation cycle and increase biofuel output. In addition, yeast concentration may be optimized to improve fermentation performance. Therefore, it is important to develop a simple method for concentration and viability measurement of fermenting yeast. In this work, we demonstrate an imaging cytometry method for concentration and viability measurements of yeast in corn mash directly from operating fermenters. It employs an automated cell counter, a dilution buffer, and staining solution from Nexcelom Bioscience to perform enumeration. The proposed method enables specific fluorescence detection of viable and nonviable yeasts, which can generate precise results for concentration and viability of yeast in corn mash. This method can provide an essential tool for research and development in the biofuel industry and may be incorporated into manufacturing to monitor yeast concentration and viability efficiently during the fermentation process.

[Onychomycosis by yeast not common in diabetics of a health center].

PubMed

Imbert, J L; G Gomez, J V; Escudero, R B; Blasco, J L

2016-10-01

Mexican diabetic population frequently presents mycosis under foot hyperkeratosis; however, in another type of onychomycosis as the ones that is assumed Candida albicans is the causal agent, it is unknown the frequency, the prevalence and if another Candida species or other yeasts are found. Evaluate the frequency of yeasts causing onychomycosis in diabetic patients looked after in public institutions of health of the State of Hidalgo, Mexico, and its association with clinical epidemiological variables. An observational, descriptive and transversal study was made on 261 patients, from which one nail sample of each one was obtained, used to isolate and identify dermatophytes and yeasts; the results were statistically correlated with 24 epidemiological parameters. The clinical study was done through interrogation and by medical exploration in order to evaluate Tinea pedis and onychomycosis. Onychomycosis were caused by Candida guilliermondii, Candida parapsilosis, Candida glabrata, Candida krusei, Candida spp., Kodamaea ohmeri, Prototheca wickerhamii and unidentified yeasts. The prevalence for general onychomycosis, by dermatophytes, mixed onychomycosis and by yeasts were: 24.1, 19.5, 2.3 and 14.6%, respectively. Patients with significant probability to be diagnosed as having onychomycosis by yeasts are those wearing open shoes (2.59%); technicians and professionals (10.49%) and alcohol drinkers (3.72%). The fact that Candida albicans is not present in this study as causal agent of onychomycosis, and emerging and non-common yeasts were indeed isolated, creates new challenges. It is remarked the clinical criterion that when onychomycosis is suspected in diabetics, the diagnosis for culturing dermatophytes and yeasts should be included. Copyright © 2015 Sociedad Española de Médicos de Atención Primaria (SEMERGEN). Publicado por Elsevier España, S.L.U. All rights reserved.

Yeast Communities of Chestnut Soils under Vineyards in Dagestan

NASA Astrophysics Data System (ADS)

Abdullabekova, D. A.; Magomedova, E. S.; Magomedov, G. G.; Aliverdieva, D. A.; Kachalkin, A. V.

2017-12-01

The study of yeast communities in chestnut soils (Kastanozems) under vineyards in the Republic of Dagestan made it possible to isolate 20 yeast species. Most of the yeasts under vineyards belonged to ascomycetes, among which species of the Saccharomycetaceae family (in particular, Saccharomyces cerevisiae) comprised a significant part. The obtained results indicate that the soils under vineyards keep the pool of microbial diversity and ensure preservation of many species typical for grapes. The method of enrichment culture on grape juice medium proved to be more efficient than other methods of analysis with respect to the number of isolated species and the rate of their detection. However, implementation of different techniques to study yeasts' diversity can give somewhat different results; a set of methods should be used for an integrated analysis.

Effects of different yeast cell wall supplements added to maize- or wheat-based diets for broiler chickens.

PubMed

Morales-López, R; Auclair, E; Van Immerseel, F; Ducatelle, R; García, F; Brufau, J

2010-06-01

1. Three experiments were carried out to study the effects of two experimental yeast cell wall (YCW) supplements, one from the yeast extract industry and the other from the brewery industry, added to maize or wheat based-diets, on performance and intestinal parameters of broiler chickens (Ross 308). 2. In the first and second experiments, a completely randomised block design with 4 experimental treatments was used: T-1) Negative control, no additives T-2) Positive control, avilamycin group (10 mg/kg feed), T-3) Yeast extract-YCW (500 mg/kg), and T-4) Brewery-YCW (500 mg/kg feed). There were 6 replicates of 20 (experiment 1) and 22 (experiment 2) chicks per treatment. 3. In experiment 1 (wheat based diets), yeast extract-YCW increased BW and daily feed intake (42 d). The effects were comparable to those of avilamycin. In experiment 2 (maize based diet), avilamycin, yeast extract-YCW and brewery-YCW treatments improved the feed conversion ratio with respect to the negative control group (0 to 14 d). 4. At 24 d, in both experiments, the ileal nutrient digestibility and ileal bacterial counts were not affected by any experimental treatment. In maize diets, lower intestinal viscosity was obtained with avilamycin, yeast extract-YCW and brewery-YCW than with the negative control. In wheat diets, yeast extract-YCW and brewery-YCW reduced intestinal viscosity. 5. A third experiment was conducted to study the effect of yeast extract-YCW on animal performance, intestinal mucosa morphology and intestinal viscosity. A 2 x 2 factorial arrangement of treatments was used; one factor was the dietary yeast extract-YCW supplementation (0 or 500 mg/kg feed) and the other the cereal in the diet (maize or wheat). 6. At 43 d, the heaviest BW was in chickens fed on yeast extract-YCW compared to those given the negative control. At 22 d, yeast extract-YCW increased villus height, mucus thickness and number of goblet cells with respect to negative control. 7. Results of these experiments suggest that supplementation of yeast extract-YCW to broiler chicken diets increased animal performance by favouring intestinal mucosal development.

Competition assays and physiological experiments of soil and phyllosphere yeasts identify Candida subhashii as a novel antagonist of filamentous fungi.

PubMed

Hilber-Bodmer, Maja; Schmid, Michael; Ahrens, Christian H; Freimoser, Florian M

2017-01-05

While recent advances in next generation sequencing technologies have enabled researchers to readily identify countless microbial species in soil, rhizosphere, and phyllosphere microbiomes, the biological functions of the majority of these species are unknown. Functional studies are therefore urgently needed in order to characterize the plethora of microorganisms that are being identified and to point out species that may be used for biotechnology or plant protection. Here, we used a dual culture assay and growth analyses to characterise yeasts (40 different isolates) and their antagonistic effect on 16 filamentous fungi; comprising plant pathogens, antagonists, and saprophytes. Overall, this competition screen of 640 pairwise combinations revealed a broad range of outcomes, ranging from small stimulatory effects of some yeasts up to a growth inhibition of more than 80% by individual species. On average, yeasts isolated from soil suppressed filamentous fungi more strongly than phyllosphere yeasts and the antagonistic activity was a species-/isolate-specific property and not dependent on the filamentous fungus a yeast was interacting with. The isolates with the strongest antagonistic activity were Metschnikowia pulcherrima, Hanseniaspora sp., Cyberlindnera sargentensis, Aureobasidium pullulans, Candida subhashii, and Pichia kluyveri. Among these, the soil yeasts (C. sargentensis, A. pullulans, C. subhashii) assimilated and/or oxidized more di-, tri- and tetrasaccharides and organic acids than yeasts from the phyllosphere. Only the two yeasts C. subhashii and M. pulcherrima were able to grow with N-acetyl-glucosamine as carbon source. The competition assays and physiological experiments described here identified known antagonists that have been implicated in the biological control of plant pathogenic fungi in the past, but also little characterised species such as C. subhashii. Overall, soil yeasts were more antagonistic and metabolically versatile than yeasts from the phyllosphere. Noteworthy was the strong antagonistic activity of the soil yeast C. subhashii, which had so far only been described from a clinical sample and not been studied with respect to biocontrol. Based on binary competition assays and growth analyses (e.g., on different carbon sources, growth in root exudates), C. subhashii was identified as a competitive and antagonistic soil yeast with potential as a novel biocontrol agent against plant pathogenic fungi.

Mechanisms of yeast stress tolerance and its manipulation for efficient fuel ethanol production.

PubMed

Zhao, X Q; Bai, F W

2009-10-12

Yeast strains of Saccharomyces cerevisiae have been extensively studied in recent years for fuel ethanol production, in which yeast cells are exposed to various stresses such as high temperature, ethanol inhibition, and osmotic pressure from product and substrate sugars as well as the inhibitory substances released from the pretreatment of lignocellulosic biomass. An in-depth understanding of the mechanism of yeast stress tolerance contributes to breeding more robust strains for ethanol production, especially under very high gravity conditions. Taking advantage of the "omics" technology, the stress response and defense mechanism of yeast cells during ethanol fermentation were further explored, and the newly emerged tools such as genome shuffling and global transcription machinery engineering have been applied to breed stress resistant yeast strains for ethanol production. In this review, the latest development of stress tolerance mechanisms was focused, and improvement of yeast stress tolerance by both random and rational tools was presented.

Nutrient supplements boost yeast transformation efficiency

PubMed Central

Yu, Sheng-Chun; Dawson, Alexander; Henderson, Alyssa C.; Lockyer, Eloise J.; Read, Emily; Sritharan, Gayathri; Ryan, Marjah; Sgroi, Mara; Ngou, Pok M.; Woodruff, Rosie; Zhang, Ruifeng; Ren Teen Chia, Travis; Liu, Yu; Xiang, Yiyu; Spanu, Pietro D.

2016-01-01

Efficiency of yeast transformation is determined by the rate of yeast endocytosis. The aim of this study was to investigate the effect of introducing amino acids and other nutrients (inositol, adenine, or p-aminobenzoic acid) in the transformation medium to develop a highly efficient yeast transformation protocol. The target of rapamycin complex 1 (TORC1) kinase signalling complex influences the rate of yeast endocytosis. TORC signaling is induced by amino acids in the media. Here, we found that increasing the concentration of amino acids and other nutrients in the growth media lead to an increase yeast transformation efficiency up to 107 CFU per μg plasmid DNA and per 108 cells with a 13.8 kb plasmid DNA. This is over 130 times that of current published methods. This improvement may facilitate more efficient experimentation in which transformation efficiency is critical, such as yeast two-hybrid screening. PMID:27760994

Yeast species diversity in apple juice for cider production evidenced by culture-based method.

PubMed

Lorenzini, Marilinda; Simonato, Barbara; Zapparoli, Giacomo

2018-05-07

Identification of yeasts isolated from apple juices of two cider houses (one located in a plain area and one in an alpine area) was carried out by culture-based method. Wallerstein Laboratory Nutrient Agar was used as medium for isolation and preliminary yeasts identification. A total of 20 species of yeasts belonging to ten different genera were identified using both BLAST algorithm for pairwise sequence comparison and phylogenetic approaches. A wide variety of non-Saccharomyces species was found. Interestingly, Candida railenensis, Candida cylindracea, Hanseniaspora meyeri, Hanseniaspora pseudoguilliermondii, and Metschnikowia sinensis were recovered for the first time in the yeast community of an apple environment. Phylogenetic analysis revealed a better resolution in identifying Metschnikowia and Moesziomyces isolates than comparative analysis using the GenBank or YeastIP gene databases. This study provides important data on yeast microbiota of apple juice and evidenced differences between two geographical cider production areas in terms of species composition.

History of genome editing in yeast.

PubMed

Fraczek, Marcin G; Naseeb, Samina; Delneri, Daniela

2018-05-01

For thousands of years humans have used the budding yeast Saccharomyces cerevisiae for the production of bread and alcohol; however, in the last 30-40 years our understanding of the yeast biology has dramatically increased, enabling us to modify its genome. Although S. cerevisiae has been the main focus of many research groups, other non-conventional yeasts have also been studied and exploited for biotechnological purposes. Our experiments and knowledge have evolved from recombination to high-throughput PCR-based transformations to highly accurate CRISPR methods in order to alter yeast traits for either research or industrial purposes. Since the release of the genome sequence of S. cerevisiae in 1996, the precise and targeted genome editing has increased significantly. In this 'Budding topic' we discuss the significant developments of genome editing in yeast, mainly focusing on Cre-loxP mediated recombination, delitto perfetto and CRISPR/Cas. © 2018 The Authors. Yeast published by John Wiley & Sons, Ltd.

Biosorption of nickel by yeasts in an osmotically unsuitable environment.

PubMed

Breierová, Emilia; Certík, Milan; Kovárová, Annamaria; Gregor, Tomas

2008-01-01

The tolerance, sorption of nickel(II) ions, and changes in the production and composition of exopolymers of eight yeast strains grown under nickel presence with/without NaCl were studied. Strains of Pichia anomala and Candida maltosa known as the most resistant yeasts against nickel tolerated up to 3 mM Ni2+. NaCl addition decreased both the resistance of the yeast strains toward nickel ions and the sorption of metal ions into cells. All yeasts absorbed nickel predominantly into exopolymers (glycoproteins) and on the surface of cells. However, while the amount of polysaccharide moieties of exoglycoproteins of most of the resistant yeasts was induced by stress conditions, the ratio polysaccharide/protein in the exopolymers remained unchanged in the sensitive species Cystofilobasidium. The exopolymer composition might play a key role in yeast adaptation to stress conditions caused by heavy metal ions.

The influence of Aster x salignus Willd. Invasion on the diversity of soil yeast communities

NASA Astrophysics Data System (ADS)

Glushakova, A. M.; Kachalkin, A. V.; Chernov, I. Yu.

2016-07-01

The annual dynamics of yeast communities were studied in the soddy-podzolic soil under the thickets of Aster x salignus Willd., one of the widespread invasive plant species in central Russia. Yeast groups in the soils under continuous aster thickets were found to differ greatly from the yeast communities in the soils under the adjacent indigenous meadow vegetation. In both biotopes the same species ( Candida vartiovaarae, Candida sake, and Cryptococcus terreus) are dominants. However, in the soils under indigenous grasses, eurybiontic yeasts Rhodotorula mucilaginosa, which almost never occur in the soil under aster, are widespread. In the soil under aster, the shares of other typical epiphytic and pedobiontic yeast fungi (ascomycetic species Wickerhamomyces aniomalus, Barnettozyma californica and basidiomycetic species Cystofilobasidium macerans, Guehomyces pullulans) significantly increase. Thus, the invasion of Aster x salignus has a clear effect on soil yeast complexes reducing their taxonomic and ecological diversity.

Micro-organisms behind the pollination scenes: microbial imprint on floral nectar sugar variation in a tropical plant community

PubMed Central

Canto, A.; Herrera, C. M.

2012-01-01

Background and Aims Variation in the composition of floral nectar reflects intrinsic plant characteristics as well as the action of extrinsic factors. Micro-organisms, particularly yeasts, represent one extrinsic factor that inhabit the nectar of animal-pollinated flowers worldwide. In this study a ‘microbial imprint hypothesis’ is formulated and tested, in which it is proposed that natural community-wide variation in nectar sugar composition will partly depend on the presence of yeasts in flowers. Methods Occurrence and density of yeasts were studied microscopically in single-flower nectar samples of 22 animal-pollinated species from coastal xeric and sub-humid tropical habitats of the Yucatán Peninsula, Mexico. Nectar sugar concentration and composition were concurrently determined on the same samples using high-performance liquid chromatography (HPLC) methods. Key Results Microscopical examination of nectar samples revealed the presence of yeasts in nearly all plant species (21 out of 22 species) and in about half of the samples examined (51·8 % of total, all species combined). Plant species and individuals differed significantly in nectar sugar concentration and composition, and also in the incidence of nectar yeasts. After statistically controlling for differences between plant species and individuals, nectar yeasts still accounted for a significant fraction of community-wide variance in all nectar sugar parameters considered. Significant yeast × species interactions on sugar parameters revealed that plant species differed in the nectar sugar correlates of variation in yeast incidence. Conclusions The results support the hypothesis that nectar yeasts impose a detectable imprint on community-wide variation in nectar sugar composition and concentration. Since nectar sugar features influence pollinator attraction and plant reproduction, future nectar studies should control for yeast presence and examine the extent to which microbial signatures on nectar characteristics ultimately have some influence on pollination services in plant communities. PMID:22915578

Development of friction material by using precast prefired (pcp f) blocks

NASA Astrophysics Data System (ADS)

Dineshkumar, R.; Ramanamurthy, E. V. V.; Krishnapavanteja, Ch

2017-05-01

The braking system used to control and stop automobile system. The braking system converts the kinematic energy into heat energy by friction. The performance of the brake pad depends on composition of friction materials. The asbestos brake pads are carcinogenic nature and it makes so many health problems. The present research work is going to replacement of asbestos by new materials. The new material is made by fused ceramic materials from industrial wastage. In this study the industrial waste are recycled and conducted the suitable test to compare the performance of the new material with existing brake pad material. The wear test was conducted by pin on disc experiment. The non asbestos, nonfused, fused samples are represented by x1, x2 and x3. The new brake pad material is formed by non fused and fused ceramic materials. The brake pads are manufactured by powder compacting process.

Contemplating the ultimate sacrifice: identity fusion channels pro-group affect, cognition, and moral decision making.

PubMed

Swann, William B; Gómez, Angel; Buhrmester, Michael D; López-Rodríguez, Lucía; Jiménez, Juan; Vázquez, Alexandra

2014-05-01

Although most people acknowledge the moral virtue in sacrificing oneself to save others, few actually endorse self-sacrifice. Seven experiments explored the cognitive and emotional mechanisms that underlie such endorsements. Participants responded to 1 of 2 moral dilemmas in which they could save 5 members of their country only by sacrificing themselves. Over 90% of participants acknowledged that the moral course of action was to sacrifice oneself to save others (Experiment 1), yet only those who were strongly fused with the group preferentially endorsed self-sacrifice (Experiments 2-7). The presence of a concern with saving group members rather than the absence of a concern with self-preservation motivated strongly fused participants to endorse sacrificing themselves for the group (Experiment 3). Analyses of think aloud protocols suggested that saving others was motivated by emotional engagement with the group among strongly fused participants but by utilitarian concerns among weakly fused participants (Experiment 4). Hurrying participants' responses increased self-sacrifice among strongly fused participants but decreased self-sacrifice among weakly fused participants (Experiment 5). Priming the personal self increased endorsement of self-sacrifice among strongly fused participants but further reduced endorsement of self-sacrifice among weakly fused participants (Experiment 6). Strongly fused participants ignored utilitarian considerations, but weakly fused persons endorsed self-sacrifice more when it would save more people (Experiment 7). Apparently, the emotional engagement with the group experienced by strongly fused persons overrides the desire for self-preservation and compels them to translate their moral beliefs into self-sacrificial behavior.

The yeast Golgi apparatus: insights and mysteries

PubMed Central

Papanikou, Effrosyni; Glick, Benjamin S.

2009-01-01

The Golgi apparatus is known to modify and sort newly synthesized secretory proteins. However, fundamental mysteries remain about the structure, operation, and dynamics of this organelle. Important insights have emerged from studying the Golgi in yeasts. For example, yeasts have provided direct evidence for Golgi cisternal maturation, a mechanism that is likely to be broadly conserved. Here, we highlight features of the yeast Golgi as well as challenges that lie ahead. PMID:19879270

Yeast: An Experimental Organism for Modern Biology.

ERIC Educational Resources Information Center

Botstein, David; Fink, Gerald R.

1988-01-01

Discusses the applicability and advantages of using yeasts as popular and ideal model systems for studying and understanding eukaryotic biology at the cellular and molecular levels. Cites experimental tractability and the cooperative tradition of the research community of yeast biologists as reasons for this success. (RT)

Quantifying the Uncertainty in High Spatial and Temporal Resolution Synthetic Land Surface Reflectance at Pixel Level Using Ground-Based Measurements

NASA Astrophysics Data System (ADS)

Kong, J.; Ryu, Y.

2017-12-01

Algorithms for fusing high temporal frequency and high spatial resolution satellite images are widely used to develop dense time-series land surface observations. While many studies have revealed that the synthesized frequent high spatial resolution images could be successfully applied in vegetation mapping and monitoring, validation and correction of fused images have not been focused than its importance. To evaluate the precision of fused image in pixel level, in-situ reflectance measurements which could account for the pixel-level heterogeneity are necessary. In this study, the synthetic images of land surface reflectance were predicted by the coarse high-frequency images acquired from MODIS and high spatial resolution images from Landsat-8 OLI using the Flexible Spatiotemporal Data Fusion (FSDAF). Ground-based reflectance was measured by JAZ Spectrometer (Ocean Optics, Dunedin, FL, USA) on rice paddy during five main growth stages in Cheorwon-gun, Republic of Korea, where the landscape heterogeneity changes through the growing season. After analyzing the spatial heterogeneity and seasonal variation of land surface reflectance based on the ground measurements, the uncertainties of the fused images were quantified at pixel level. Finally, this relationship was applied to correct the fused reflectance images and build the seasonal time series of rice paddy surface reflectance. This dataset could be significant for rice planting area extraction, phenological stages detection, and variables estimation.

Synthesis, reactivity, and properties of N-fused porphyrin rhenium(I) tricarbonyl complexes.

PubMed

Toganoh, Motoki; Ikeda, Shinya; Furuta, Hiroyuki

2007-11-12

The thermal reactions of N-fused tetraarylporphyrins or N-confused tetraarylporphyrins with Re2(CO)10 gave the rhenium(I) tricarbonyl complexes bearing N-fused porphyrinato ligands (4) in moderate to good yields. The rhenium complexes 4 are characterized by mass, IR, 1H, and 13C NMR spectroscopy, and the structures of tetraphenylporphynato complex 4a and its nitro derivative 15 are determined by X-ray single crystal analysis. The rhenium complexes 4 show excellent stability against heat, light, acids, bases, and oxidants. The aromatic substitution reactions of 4 proceed without a loss of the center metal to give the nitro (15), formyl (16), benzoyl (17), and cyano derivatives (19), regioselectively. In the electrochemical measurements for 4, one reversible oxidation wave and two reversible reduction waves are observed. Their redox potentials imply narrow HOMO-LUMO band gaps of 4 and are consistent with their electronic absorption spectra, in which the absorption edges exceed 1000 nm. Theoretical study reveals that the HOMO and LUMO of the rhenium complexes are exclusively composed of the N-fused porphyrin skeleton. Protonation of 4 takes place at the 21-position regioselectively, reflecting the high coefficient of the C21 atom in the HOMO orbital. The skeletal rearrangement reaction from N-confused porphyrin Re(I) complex (8) to N-fused porphyrin Re(I) complex (4) is suggested from the mechanistic study as well as DFT calculations.

Production of astaxanthin rich feed supplement for animals from Phaffia rhodozyma yeast at low cost

NASA Astrophysics Data System (ADS)

Irtiza, Ayesha; Shatunova, Svetlana; Glukhareva, Tatiana; Kovaleva, Elena

2017-09-01

Dietary nutrients such as amino acids, vitamins, minerals and antioxidants can play a significant role in determining meat quality and also the growth rate of poultry or animal. Phaffia rhodozyma was grown on waste from brewery industry to produce astaxanthin rich feed supplements at a very low cost. Phaffia rhodozyma is yeast specie that has ability to produce carotenoids and approximately 80% of its total carotenoid content is astaxanthin, which is highly valuable carotenoid for food, feed and aquaculture industry. This study was carried out to test yeast extract of spent yeast from brewing industry waste (residual yeast) as potential nitrogen source for growth of Phaffia rhodozyma. Cultivation was carried out in liquid media prepared by yeast extracts and other components (glucose and peptone). Carotenoids from the biomass were released into biomass by suspending cells in DMSO for destruction of cells followed by extraction with petroleum ether. The extracted carotenoids were studied by spectrophotometry to identify and quantify astaxanthin and other carotenoids produced.

Stress tolerance and biocontrol performance of the yeast antagonist, Candida diversa, change with morphology transition.

PubMed

Li, Guangkun; Chi, Mengshan; Chen, Huizhen; Sui, Yuan; Li, Yan; Liu, Yongsheng; Zhang, Xiaojing; Sun, Zhiqiang; Liu, Guoqing; Wang, Qi; Liu, Jia

2016-02-01

As an eco-friendly management method, biological control of postharvest diseases, utilizing antagonistic yeasts, is a research topic receiving considerable attention. Detailed knowledge on the biology of yeast antagonists is crucial when considering their potential application and development as biocontrol products. Changes in the growth form, such as single-cell to pseudohyphae, have been associated with the mode of action in postharvest biocontrol yeasts. In this study, the antagonistic yeast, Candida diversa, reversibly shifted from a single-cell morphology on yeast peptone dextrose (YPD) medium with 2 % agar to a pseudohyphal morphology on YPD with 0.3 % agar. The tolerance of the pseudohyphal form to heat and oxidative stresses, as well as the biocontrol efficacy against Botrytis cinerea on apple and kiwifruit stored at 25 and 4 °C, was significantly higher as compared to the single-cell form. This study provides new information on the ability of C. diversa to change its morphology and the impact of the morphology shift on stress tolerance and biocontrol performance.

Elicitation effect of Saccharomyces cerevisiae yeast extract on main health-promoting compounds and antioxidant and anti-inflammatory potential of butter lettuce (Lactuca sativa L.).

PubMed

Złotek, Urszula; Świeca, Michał

2016-05-01

This paper presents a study on changes in the main phytochemical levels and antioxidant and anti-inflammatory activity of lettuce caused by different doses and times of application of yeast extracts. Elicitation with yeast extract caused an increase in the total phenolic compounds and chlorophyll content, which varied according to the dose and time of spraying, but it did not have a positive impact on vitamin C, flavonoid and carotenoid content in lettuce. The best effect was achieved by double spraying with 1% yeast extract and by single spraying with 0.1% yeast extract. The increase in phytochemical content was positively correlated with the antioxidant and anti-inflammatory activity of the studied lettuce leaves. Chicoric acid seems to be the major contributor to these antioxidant activities. Yeast extract may be used as a natural, environmentally friendly and safe elicitor for improving the health-promoting qualities of lettuce. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

Diversity of culturable yeasts associated with zoanthids from Brazilian reef and its relation with anthropogenic disturbance.

PubMed

Paulino, Gustavo Vasconcelos Bastos; Félix, Ciro Ramon; Broetto, Leonardo; Landell, Melissa Fontes

2017-10-15

Some of the main threats to coral reefs come from human actions on marine environment, such as tourism, overfishing and pollution from urban development. While several studies have demonstrated an association between bacteria and corals, demonstrating how these communities react to different anthropogenic stressors, yeast communities associated with corals have received far less attention from researchers. The aim of this work was therefore to describe cultivable yeasts associated with three coral species and to evaluate the influence of sewage discharge on yeasts community. We obtained 130 isolates, mostly belonging to phylum Ascomycota and many of them had previously been isolated from human samples or are considered pathogens. The mycobiota was more similar among corals collected from the same reef, indicating that the composition of reef yeast community is more influenced by environmental conditions than host species. We suggest further studies to elucidate which factors are most influential on the composition of the coral-associated yeast community. Copyright © 2017 Elsevier Ltd. All rights reserved.

16 CFR 1507.3 - Fuses.

Code of Federal Regulations, 2010 CFR

2010-01-01

... Practices CONSUMER PRODUCT SAFETY COMMISSION FEDERAL HAZARDOUS SUBSTANCES ACT REGULATIONS FIREWORKS DEVICES § 1507.3 Fuses. (a) Fireworks devices that require a fuse shall: (1) Utilize only a fuse that has been... it will support either the weight of the fireworks device plus 8 ounces of dead weight or double the...

FUSE: a profit maximization approach for functional summarization of biological networks.

PubMed

Seah, Boon-Siew; Bhowmick, Sourav S; Dewey, C Forbes; Yu, Hanry

2012-03-21

The availability of large-scale curated protein interaction datasets has given rise to the opportunity to investigate higher level organization and modularity within the protein interaction network (PPI) using graph theoretic analysis. Despite the recent progress, systems level analysis of PPIS remains a daunting task as it is challenging to make sense out of the deluge of high-dimensional interaction data. Specifically, techniques that automatically abstract and summarize PPIS at multiple resolutions to provide high level views of its functional landscape are still lacking. We present a novel data-driven and generic algorithm called FUSE (Functional Summary Generator) that generates functional maps of a PPI at different levels of organization, from broad process-process level interactions to in-depth complex-complex level interactions, through a pro t maximization approach that exploits Minimum Description Length (MDL) principle to maximize information gain of the summary graph while satisfying the level of detail constraint. We evaluate the performance of FUSE on several real-world PPIS. We also compare FUSE to state-of-the-art graph clustering methods with GO term enrichment by constructing the biological process landscape of the PPIS. Using AD network as our case study, we further demonstrate the ability of FUSE to quickly summarize the network and identify many different processes and complexes that regulate it. Finally, we study the higher-order connectivity of the human PPI. By simultaneously evaluating interaction and annotation data, FUSE abstracts higher-order interaction maps by reducing the details of the underlying PPI to form a functional summary graph of interconnected functional clusters. Our results demonstrate its effectiveness and superiority over state-of-the-art graph clustering methods with GO term enrichment.

Unraveling the Enzymatic Basis of Wine “Flavorome”: A Phylo-Functional Study of Wine Related Yeast Species

PubMed Central

Belda, Ignacio; Ruiz, Javier; Alastruey-Izquierdo, Ana; Navascués, Eva; Marquina, Domingo; Santos, Antonio

2016-01-01

Non-Saccharomyces yeasts are a heterogeneous microbial group involved in the early stages of wine fermentation. The high enzymatic potential of these yeasts makes them a useful tool for increasing the final organoleptic characteristics of wines in spite of their low fermentative power. Their physiology and contribution to wine quality are still poorly understood, with most current knowledge being acquired empirically and in most cases based in single species and strains. This work analyzed the metabolic potential of 770 yeast isolates from different enological origins and representing 15 different species, by studying their production of enzymes of enological interest and linking phylogenetic and enzymatic data. The isolates were screened for glycosidase enzymes related to terpene aroma release, the β-lyase activity responsible for the release of volatile thiols, and sulfite reductase. Apart from these aroma-related activities, protease, polygalacturonase and cellulase activities were also studied in the entire yeast collection, being related to the improvement of different technological and sensorial features of wines. In this context, and in terms of abundance, two different groups were established, with α-L-arabinofuranosidase, polygalacturonase and cellulase being the less abundant activities. By contrast, β-glucosidase and protease activities were widespread in the yeast collection studied. A classical phylogenetic study involving the partial sequencing of 26S rDNA was conducted in conjunction with the enzymatic profiles of the 770 yeast isolates for further typing, complementing the phylogenetic relationships established by using 26S rDNA. This has rendered it possible to foresee the contribution different yeast species make to wine quality and their potential applicability as pure inocula, establishing species-specific behavior. These consistent results allowed us to design future targeted studies on the impact different non-Saccharomyces yeast species have on wine quality, understanding intra and interspecific enzymatic odds and, therefore, aiming to predict the most suitable application for the current non-Saccharomyces strains, as well as the potential future applications of new strains. This work therefore contributes to a better understanding of the concept of wine microbiome and its potential consequences for wine quality, as well as to the knowledge of non-Saccharomyces yeasts for their use in the wine industry. PMID:26834730

Enological Qualities and Interactions Between Native Yeast and Lactic Acid Bacteria from Queretaro, Mexico.

PubMed

Miranda-Castilleja, Dalia E; Martínez-Peniche, Ramón Á; Nadal Roquet-Jalmar, Montserrat; Aldrete-Tapia, J Alejandro; Arvizu-Medrano, Sofía M

2018-06-15

Despite the importance of strain compatibility, most of the enological strain selection studies are carried out separately on yeasts and lactic acid bacteria (LAB). In this study, the enological traits and interactions between native yeasts and LAB were studied. The H 2 S and acetic acid production, growth rates at 8 °C, killer phenotypes, flocculation, and tolerance to must and wine inhibitors were determined for 25 Saccharomyces yeasts. The ability to grow under two wine-like conditions was also determined in 37 LAB (Oenococcus oeni and Lactobacillus plantarum). The yeast-LAB compatibility of selected strains was tested in a sequential scheme. Finally, microvinification trials were performed using two strains from each group to determine the efficiencies and quality parameters. The phenotypic characterization by the K-means and hierarchical clusters indicated a correlation between flocculation and optical density increase in simulated must and wine medium (r = -0.415) and grouped the prominent yeasts SR19, SR26, and N05 as moderately flocculent, killer, acid producing, and highly tolerant strains. Among the LAB, L. plantarum FU39 grew 230% more than the rest. With regard to interactions, LAB growth stimulation (14-fold on average) due to the previous action of yeasts, particularly of SR19, was observed. The final quality of all wines was similar, but yeast SR19 performed a faster and more efficient fermentation than did N05, Also L. plantarum FU39 fermented faster than did O. oeni VC32. The use of quantitative data, and multivariate analyses allowed an integrative approach to the selection of a compatible and efficient pair of enological yeast-LAB strains. An alternative scheme is proposed for the joint selection of yeast and lactic acid bacteria strains, which allows us to foresee the interactions that may occur between them during winemaking. The kinetic parameters, turbidimetrically measured and analyzed by multivariate methods, simplify the detection of outstanding selectable microorganisms. This methodology can be implemented at any cellar or even any fermentative industry that aims to select compatible yeast and lactic acid bacteria. © 2018 Institute of Food Technologists®.

Effect of processing parameters on surface finish for fused deposition machinable wax patterns

NASA Technical Reports Server (NTRS)

Roberts, F. E., III

1995-01-01

This report presents a study on the effect of material processing parameters used in layer-by-layer material construction on the surface finish of a model to be used as an investment casting pattern. The data presented relate specifically to fused deposition modeling using a machinable wax.

Tracking of "Moving" Fused Auditory Images by Children.

ERIC Educational Resources Information Center

Cranford, Jerry L.; And Others

1993-01-01

This study evaluated the ability of 30 normally developing children (ages 6-12) to report the perceived location of a stationary fused auditory image (FAI) or track a "moving" FAI. Although subjects performed at normal adult levels with the stationary sound measure, they exhibited a significant age-related trend with the moving sound…

Studies on transmitted beam modulation effect from laser induced damage on fused silica optics.

PubMed

Zheng, Yi; Ma, Ping; Li, Haibo; Liu, Zhichao; Chen, Songlin

2013-07-15

UV laser induced damage (LID) on exit surface of fused silica could cause modulation effect to transmitted beam and further influence downstream propagation properties. This paper presents our experimental and analytical studies on this topic. In experiment, a series of measurement instruments are applied, including beam profiler, interferometer, microscope, and optical coherent tomography (OCT). Creating and characterizing of LID on fused silica sample have been implemented. Morphological features are studied based on their particular modulation effects on transmitted beam. In theoretical investigation, analytical modeling and numerical simulation are performed. Modulation effects from amplitude, phase, and size factors are analyzed respectively. Furthermore, we have novelly designed a simplified polygon model to simulate actual damage site with multiform modulation features, and the simulation results demonstrate that the modeling is usable and representative.

Gut yeast communities in Larus michahellis from various breeding colonies.

PubMed

Al-Yasiri, Mohammed Hashim; Normand, Anne-Cécile; Piarroux, Renaud; Ranque, Stéphane; Mauffrey, Jean-François

2017-06-01

Yellow-legged gulls have been reported to carry antibiotic-resistant Enterobacteriaceae; however, the gut mycobiota of these birds has not yet been described. In this study, we analyzed the gut yeast communities in five yellow-legged gull breeding colonies along the Mediterranean littoral in southern France. Gull fecal samples were inoculated onto four types of culture media, including one supplemented with itraconazole. Yeast species richness, abundance, and diversity were estimated, and factorial analysis was used to highlight correspondences between breeding colonies. Yeast grew in 113 of 177 cultures, and 17 distinct yeast species were identified. The most frequent species were Candida krusei (53.5%), Galactomyces geotrichum (44.1%), C. glabrata (40.9%), C. albicans (20.5%), and Saccharomyces cerevisiae (18.1%). Gut yeast community structure in the gulls at both Pierre-Blanche Lagoon (PB) and Frioul Archipelago (F) were characterized by greater species richness and diversity than in those at the two cities of La Grande-Motte (GM) and Palavas-les-Flots (PF) as well as Riou Archipelago (R). Gulls in these latter three sites probably share a similar type of anthropogenic diet. Notably, the proportion of anthropic yeast species, including C. albicans and C. glabrata, in the gull mycobiota increased with gull colony synanthropy. Antifungal resistance was found in each of the five most frequent yeast species. We found that the gut yeast communities of these yellow-legged gulls include antifungal-resistant human pathogens. Further studies should assess the public health impact of these common synanthropic seabirds, which represent a reservoir and disseminator of drug-resistant human pathogenic yeast into the environment. © The Author 2016. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Accountability for Information Flow via Explicit Formal Proof

DTIC Science & Technology

2009-10-01

macrobenchmarks. The first (called OpenSSL in the table below), unpacks the OpenSSL source code, compiles it and deletes it. The other (called Fuse in...penalty for PCFS as compared to Fuse/Null is approximately 10% for OpenSSL , and 2.5% for Fuse. The difference arises because the OpenSSL benchmark depends...Macrobenchmarks Benchmark PCFS Fuse/Null Ext3 OpenSSL 126 114 94 Fuse x 5 79 77 70 15 In summary, assuming a low rate of cache misses, the

Seasonal and plant-dependent variations in diversity, abundance and stress tolerance of epiphytic yeasts in desert habitats.

PubMed

Abu-Ghosh, Said; Droby, Samir; Korine, Carmi

2014-08-01

We studied the epiphytic yeast species of the plants of the Negev Desert and the Dead Sea region, Israel, which are considered one of the most extreme hyper-arid lands in the world. For this purpose, we developed isolation protocols; we performed morphological, cultural and molecular identification tests and compared yeast diversity between the locations and the plants. The composition of the yeast populations present in the study's plants underwent seasonal fluctuations, whereas differences in community compositions were significant within sites. The maximum number of species of yeast occurred in autumn and Cryptococcus spp. were predominant year round. The isolated yeast strains showed an unusual tolerance to extreme growth conditions, such as high temperatures (up to 72% viability at 50°C), lethal hydrogen peroxide and NaCl concentrations. These results suggest that epiphytic yeasts inhabit the plants of the Dead Sea region and the Negev Desert have a community structure that is unique to the plant species and have a high tolerance to the harsh conditions that enables them to adapt to an arid ecosystem. © 2014 Society for Applied Microbiology and John Wiley & Sons Ltd.

Characterization of butter spoiling yeasts and their inhibition by some spices.

PubMed

Sagdic, Osman; Ozturk, Ismet; Bayram, Okan; Kesmen, Zulal; Yilmaz, Mustafa Tahsin

2010-01-01

This study was designed to identify the yeasts in packaged and unpackaged butters and screen antiyeast activity of spices, including marjoram (Origanum majorana L.), summer savory (Satureja hortensis L.), and black cumin (Nigella sativa L.) against the most dominant yeast species in the packaged and unpackaged butters. Mean total yeast populations were 5.40 log CFU/g in unpackaged butter samples and 2.22 log CFU/g in packaged butter samples, indicating better hygienic quality of packaged samples. Forty-nine yeast species were isolated and identified from butter samples with the most prevalent isolates belonging to genera Candida-C. kefyr, C. zeylanoides, and C. lambica-and with moderate number of isolates belonging to genera Cryptococcus, Rhodotorula, Saccharomyces, and Zygosaccharomyces. Black cumin exhibited the highest antiyeast activity against C. zeylanoides and C. lambica species, even inhibited these species, while summer savory inhibited C. kefyr. The results of this study revealed clear antimicrobial potential of black cumin against the yeast species isolated from butters. Marjoram, summer savory, and black cumin could be used as natural antimicrobial agents against spoilage yeasts in food preservation, especially in butter. © 2010 Institute of Food Technologists®

Differential Adsorption of Ochratoxin A and Anthocyanins by Inactivated Yeasts and Yeast Cell Walls during Simulation of Wine Aging

PubMed Central

Petruzzi, Leonardo; Baiano, Antonietta; De Gianni, Antonio; Sinigaglia, Milena; Corbo, Maria Rosaria; Bevilacqua, Antonio

2015-01-01

The adsorption of ochratoxin A (OTA) by yeasts is a promising approach for the decontamination of musts and wines, but some potential competitive or interactive phenomena between mycotoxin, yeast cells, and anthocyanins might modify the intensity of the phenomenon. The aim of this study was to examine OTA adsorption by two strains of Saccharomyces cerevisiae (the wild strain W13, and the commercial isolate BM45), previously inactivated by heat, and a yeast cell wall preparation. Experiments were conducted using Nero di Troia red wine contaminated with 2 μg/L OTA and supplemented with yeast biomass (20 g/L). The samples were analyzed periodically to assess mycotoxin concentration, chromatic characteristics, and total anthocyanins over 84 days of aging. Yeast cell walls revealed the highest OTA-adsorption in comparison to thermally-inactivated cells (50% vs. 43% toxin reduction), whilst no significant differences were found for the amount of adsorbed anthocyanins in OTA-contaminated and control wines. OTA and anthocyanins adsorption were not competitive phenomena. Unfortunately, the addition of yeast cells to wine could cause color loss; therefore, yeast selection should also focus on this trait to select the best strain. PMID:26516913

Single nucleotide polymorphisms of PAD1 and FDC1 show a positive relationship with ferulic acid decarboxylation ability among industrial yeasts used in alcoholic beverage production.

PubMed

Mukai, Nobuhiko; Masaki, Kazuo; Fujii, Tsutomu; Iefuji, Haruyuki

2014-07-01

Among industrial yeasts used for alcoholic beverage production, most wine and weizen beer yeasts decarboxylate ferulic acid to 4-vinylguaiacol, which has a smoke-like flavor, whereas sake, shochu, top-fermenting, and bottom-fermenting yeast strains lack this ability. However, the factors underlying this difference among industrial yeasts are not clear. We previously confirmed that both PAD1 (phenylacrylic acid decarboxylase gene, YDR538W) and FDC1 (ferulic acid decarboxylase gene, YDR539W) are essential for the decarboxylation of phenylacrylic acids in Saccharomyces cerevisiae. In the present study, single nucleotide polymorphisms (SNPs) of PAD1 and FDC1 in sake, shochu, wine, weizen, top-fermenting, bottom-fermenting, and laboratory yeast strains were examined to clarify the differences in ferulic acid decarboxylation ability between these types of yeast. For PAD1, a nonsense mutation was observed in the gene sequence of standard top-fermenting yeast. Gene sequence analysis of FDC1 revealed that sake, shochu, and standard top-fermenting yeasts contained a nonsense mutation, whereas a frameshift mutation was identified in the FDC1 gene of bottom-fermenting yeast. No nonsense or frameshift mutations were detected in laboratory, wine, or weizen beer yeast strains. When FDC1 was introduced into sake and shochu yeast strains, the transformants exhibited ferulic acid decarboxylation activity. Our findings indicate that a positive relationship exists between SNPs in PAD1 and FDC1 genes and the ferulic acid decarboxylation ability of industrial yeast strains. Copyright © 2013 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Surface Aggregation of Candida albicans on Glass in the Absence and Presence of Adhering Streptococcus gordonii in a Parallel-Plate Flow Chamber: A Surface Thermodynamical Analysis Based on Acid-Base Interactions.

PubMed

Millsap; Bos; Busscher; van der Mei HC

1999-04-15

Adhesive interactions between yeasts and bacteria are important in the maintenance of infectious mixed biofilms on natural and biomaterial surfaces in the human body. In this study, the extended DLVO (Derjaguin-Landau-Verwey-Overbeek) approach has been applied to explain adhesive interactions between C. albicans ATCC 10261 and S. gordonii NCTC 7869 adhering on glass. Contact angles with different liquids and the zeta potentials of both the yeasts and bacteria were determined and their adhesive interactions were measured in a parallel-plate flow chamber.Streptococci were first allowed to adhere to the bottom glass plate of the flow chamber to different seeding densities, and subsequently deposition of yeasts was monitored with an image analysis system, yielding the degree of initial surface aggregation of the adhering yeasts and their spatial arrangement in a stationary end point. Irrespective of growth temperature, the yeast cells appeared uncharged in TNMC buffer, but yeasts grown at 37 degrees C were intrinsically more hydrophilic and had an increased electron-donating character than cells grown at 30 degrees C. All yeasts showed surface aggregation due to attractive Lifshitz-van der Waals forces. In addition, acid-base interactions between yeasts, yeasts and the glass substratum, and yeasts and the streptococci were attractive for yeasts grown at 30 degrees C, but yeasts grown at 37 degrees C only had favorable acid-base interactions with the bacteria, explaining the positive relationship between the surface coverage of the glass by streptococci and the surface aggregation of the yeasts. Copyright 1999 Academic Press.

Central Stars of Planetary Nebulae in the LMC

NASA Technical Reports Server (NTRS)

Bianchi, Luciana

2004-01-01

In FUSE cycle 2's program B001 we studied Central Stars of Planetary Nebulae (CSPN) in the Large Magellanic Could. All FUSE observations have been successfully completed and have been reduced, analyzed and published. The analysis and the results are summarized below. The FUSE data were reduced using the latest available version of the FUSE calibration pipeline (CALFUSE v2.2.2). The flux of these LMC post-AGB objects is at the threshold of FUSE's sensitivity, and thus special care in the background subtraction was needed during the reduction. Because of their faintness, the targets required many orbit-long exposures, each of which typically had low (target) count-rates. Each calibrated extracted sequence was checked for unacceptable count-rate variations (a sign of detector drift), misplaced extraction windows, and other anomalies. All the good calibrated exposures were combined using FUSE pipeline routines. The default FUSE pipeline attempts to model the background measured off-target and subtracts it from the target spectrum. We found that, for these faint objects, the background appeared to be over-estimated by this method, particularly at shorter wavelengths (i.e., < 1000 A). We therefore tried two other reductions. In the first method, subtraction of the measured background is turned off and and the background is taken to be the model scattered-light scaled by the exposure time. In the second one, the first few steps of the pipeline were run on the individual exposures (correcting for effects unique to each exposure such as Doppler shift, grating motions, etc). Then the photon lists from the individual exposures were combined, and the remaining steps of the pipeline run on the combined file. Thus, more total counts for both the target and background allowed for a better extraction.

Fracture and Fatigue Resistance of Cemented versus Fused CAD-on Veneers over Customized Zirconia Implant Abutments.

PubMed

Nossair, Shereen Ahmed; Aboushelib, Moustafa N; Morsi, Tarek Salah

2015-01-05

To evaluate the fracture mechanics of cemented versus fused CAD-on veneers on customized zirconia implant abutments. Forty-five identical customized CAD/CAM zirconia implant abutments (0.5 mm thick) were prepared and seated on short titanium implant abutments (Ti base). A second scan was made to fabricate 45 CAD-on veneers (IPS Empress CAD, A2). Fifteen CAD-on veneers were cemented on the zirconia abutments (Panavia F2.0). Another 15 were fused to the zirconia abutments using low-fusing glass, while manually layered veneers served as control (n = 15). The restorations were subjected to artificial aging (3.2 million cycles between 5 and 10 kg in a water bath at 37°C) before being axially loaded to failure. Fractured specimens were examined using scanning electron microscopy to detect fracture origin, location, and size of critical crack. Stress at failure was calculated using fractography principles (alpha = 0.05). Cemented CAD-on restorations demonstrated significantly higher (F = 72, p < 0.001) fracture load compared to fused CAD-on and manually layered restorations. Fractographic analysis of fractured specimens indicated that cemented CAD-on veneers failed due to radial cracks originating from the veneer/resin interface. Branching of the critical crack was observed in the bulk of the veneer. Fused CAD-on veneers demonstrated cohesive fracture originating at the thickest part of the veneer ceramic, while manually layered veneers failed due to interfacial fracture at the zirconia/veneer interface. Within the limitations of this study, cemented CAD-on veneers on customized zirconia implant abutments demonstrated higher fracture than fused and manually layered veneers. © 2014 by the American College of Prosthodontists.

Hydrolysis of polycarbonate in sub-critical water in fused silica capillary reactor with in situ Raman spectroscopy

USGS Publications Warehouse

Pan, Z.; Chou, I-Ming; Burruss, R.C.

2009-01-01

The advantages of using fused silica capillary reactor (FSCR) instead of conventional autoclave for studying chemical reactions at elevated pressure and temperature conditions were demonstrated in this study, including the allowance for visual observation under a microscope and in situ Raman spectroscopic characterization of polycarbonate and coexisting phases during hydrolysis in subcritical water.

Comparison between MALDI-TOF MS and FilmArray Blood Culture Identification panel for rapid identification of yeast from positive blood culture.

PubMed

Paolucci, M; Foschi, C; Tamburini, M V; Ambretti, S; Lazzarotto, T; Landini, M P

2014-09-01

In this study we evaluated MALDI-TOF MS and FilmArray methods for the rapid identification of yeast from positive blood cultures. FilmArray correctly identified 20/22 of yeast species, while MALDI-TOF MS identified 9/22. FilmArray is a reliable and rapid identification system for the direct identification of yeasts from positive blood cultures. Copyright © 2014 Elsevier B.V. All rights reserved.

Everything You Ever Wanted to Know About Saccharomyces cerevisiae Telomeres: Beginning to End

PubMed Central

Zakian, Virginia A.

2012-01-01

The mechanisms that maintain the stability of chromosome ends have broad impact on genome integrity in all eukaryotes. Budding yeast is a premier organism for telomere studies. Many fundamental concepts of telomere and telomerase function were first established in yeast and then extended to other organisms. We present a comprehensive review of yeast telomere biology that covers capping, replication, recombination, and transcription. We think of it as yeast telomeres—soup to nuts. PMID:22879408

Antioxidant N-acetyltransferase Mpr1/2 of industrial baker's yeast enhances fermentation ability after air-drying stress in bread dough.

PubMed

Sasano, Yu; Takahashi, Shunsuke; Shima, Jun; Takagi, Hiroshi

2010-03-31

During bread-making processes, yeast cells are exposed to multiple stresses. Air-drying stress is one of the most harmful stresses by generation of reactive oxygen species (ROS). Previously, we discovered that the novel N-acetyltransferase Mpr1/2 confers oxidative stress tolerance by reducing intracellular ROS level in Saccharomyces cerevisiae Sigma1278b strain. In this study, we revealed that Japanese industrial baker's yeast possesses one MPR gene. The nucleotide sequence of the MPR gene in industrial baker's yeast was identical to the MPR2 gene in Sigma1278b strain. Gene disruption analysis showed that the MPR2 gene in industrial baker's yeast is involved in air-drying stress tolerance by reducing the intracellular oxidation levels. We also found that expression of the Lys63Arg and Phe65Leu variants with enhanced enzymatic activity and stability, respectively, increased the fermentation ability of bread dough after exposure to air-drying stress compared with the wild-type Mpr1. In addition, our recent study showed that industrial baker's yeast cells accumulating proline exhibited enhanced freeze tolerance in bread dough. Proline accumulation also enhanced the fermentation ability after air-drying stress treatment in industrial baker's yeast. Hence, the antioxidant enzyme Mpr1/2 could be promising for breeding novel yeast strains that are tolerant to air-drying stress. Copyright 2010 Elsevier B.V. All rights reserved.

Yeast derived from lignocellulosic biomass as a sustainable feed resource for use in aquaculture.

PubMed

Øverland, Margareth; Skrede, Anders

2017-02-01

The global expansion in aquaculture production implies an emerging need of suitable and sustainable protein sources. Currently, the fish feed industry is dependent on high-quality protein sources of marine and plant origin. Yeast derived from processing of low-value and non-food lignocellulosic biomass is a potential sustainable source of protein in fish diets. Following enzymatic hydrolysis, the hexose and pentose sugars of lignocellulosic substrates and supplementary nutrients can be converted into protein-rich yeast biomass by fermentation. Studies have shown that yeasts such as Saccharomyces cerevisiae, Candida utilis and Kluyveromyces marxianus have favourable amino acid composition and excellent properties as protein sources in diets for fish, including carnivorous species such as Atlantic salmon and rainbow trout. Suitable downstream processing of the biomass to disrupt cell walls is required to secure high nutrient digestibility. A number of studies have shown various immunological and health benefits from feeding fish low levels of yeast and yeast-derived cell wall fractions. This review summarises current literature on the potential of yeast from lignocellulosic biomass as an alternative protein source for the aquaculture industry. It is concluded that further research and development within yeast production can be important to secure the future sustainability and economic viability of intensive aquaculture. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

Advantages of Data Fusion: First Multivariate Curve Resolution Analysis of Fused Liquid Chromatographic Second-Order Data with Dual Diode Array-Fluorescent Detection.

PubMed

Pellegrino Vidal, Rocío B; Ibañez, Gabriela A; Escandar, Graciela M

2017-03-07

For the first time, liquid chromatography-diode array detection (LC-DAD) and liquid-chromatography fluorescence detection (LC-FLD) second-order data, collected in a single chromatographic run, were fused and chemometrically processed for the quantitation of coeluting analytes. Two different experimental mixtures composed of fluorescent and nonfluorescent endocrine disruptors were analyzed. Adequate pretreatment of the matrices before their fusion was crucial to attain reliable results. Multivariate curve resolution-alternating least-squares (MCR-ALS) was applied to LC-DAD, LC-FLD, and fused LC-DAD-FLD data. Although different degrees of improvement are observed when comparing the fused matrix results in relation to those obtained using a single detector, clear benefits of data fusion are demonstrated through: (1) the obtained limits of detection in the ranges 2.1-24 ng mL -1 and 0.9-6.3 ng mL -1 for the two evaluated systems and (2) the low relative prediction errors, below 7% in all cases, indicating good recoveries and precision. The feasibility of fusing data and its advantages in the analysis of real samples was successfully assessed through the study of spiked tap, underground, and river water samples.

Vegemite Beer: yeast extract spreads as nutrient supplements to promote fermentation.

PubMed

Kerr, Edward D; Schulz, Benjamin L

2016-01-01

Vegemite is an iconic Australian food spread made from spent brewers' yeast extract, which has been reported to be used as an ingredient in illegal home brewing. In this study, we tested the utility of Vegemite and the similar spread Marmite in promoting fermentation. We could not culture microorganisms from either Vegemite or Marmite, consistent with these food-grade spreads being essentially sterile. To test if the addition of Vegemite or Marmite could assist in fermentation when additional viable yeast was also present, solutions containing glucose and a range of concentrations of either Vegemite or Marmite were inoculated with brewers' yeast. No fermentation occurred in any condition without addition of extra brewer's yeast. Fermentation did not occur when yeast was inoculated into solutions containing only glucose, but progressed efficiently with when Vegemite or Marmite was also added. Gas Chromatography confirmed that ethanol was present at ∼3% v/v post-fermentation in all samples which contained glucose, Vegemite or Marmite, and brewers' yeast. Trace amounts of methanol were also detected. Mass spectrometry proteomics identified abundant intracellular yeast proteins and barley proteins in Vegemite and Marmite, and abundant secreted yeast proteins from actively growing yeast in those samples to which extra brewers' yeast had been added. We estimate that the real-world cost of home brewed "Vegemite Beer" would be very low. Our results show that Vegemite or other yeast extract spreads could provide cheap and readily available sources of nutrient supplementation to increase the efficiency of fermentation in home brewing or other settings.

The presence of a mycangium in European Sinodendron cylindricum (Coleoptera: Lucanidae) and the associated yeast symbionts

PubMed Central

Tanahashi, Masahiko; Hawes, Colin J.

2016-01-01

Part of the exoskeleton of some wood-inhabiting insects is modified to form a mycangium, which is a specialized organ used to convey fungal spores or yeasts to their offspring. Although most stag beetles (Coleoptera: Lucanidae) are known to have female-specific mycangia and associated yeast symbionts, the evolutionary origin of the mycangium in this group remains unresolved. Here, we report the presence of a mycangium and associated yeast symbionts in the European horned stag beetle Sinodendron cylindricum (L.), which belongs to an ancestral clade of the Lucanidae. The mycangium of S. cylindricum is shown to be female-specific and have the same developmental origin as that of other stag beetles. A total of five yeast strains were isolated from adult mycangia and larval gut of S. cylindricum. Of these, we suggest that SICYAM1 is an undescribed yeast with taxonomic novelty, and have identified SICYLG3 as the xylose-fermenting yeast Scheffersomyces insectosa using nuclear ribosomal RNA and ITS sequences. The remaining three yeast strains, SICYAM2, SICYLG1, and SICYLG2, were assigned to the genus Sugiyamaella. Yeast density in the adult mycangium was lower than that of the more evolutionarily advanced stag beetles, the European Lucanus cervus (L.) and Dorcus parallelipipedus (L.), which were also examined in this study. No living yeasts were isolated from the adult guts. However, a third instar larva of S. cylindricum harbored 104–106 living yeasts in each gut region, which suggests that gut yeasts play an important role in these wood-feeding larvae. PMID:27432353

Study of the counts, species and characteristics of the yeast population during the manufacture of dry-cured "lacón". Effect of salt level.

PubMed

Purriños, Laura; García Fontán, María C; Carballo, Javier; Lorenzo, José M

2013-05-01

The aim of this work was to study the yeast population during the manufacture of dry-cured "lacón" (a Spanish traditional meat product) and the effect of the salting time. For this study, six batches of "lacón" were manufactured with three different salting times (LS (3 days of salting), MS (4 days of salting) and HS (5 days of salting)). Yeast counts increased significantly (P < 0.001) during the whole process from 2.60 to 6.37 log cfu/g. An increased length of salting time did not affect yeast counts throughout the manufacture of dry-cured "lacón", although the highest yeast counts were obtained from LS batches. A total of 226 isolates were obtained from dry-cured "lacón" during drying-ripening stage, of which 151 were yeasts and were identified at the species level using molecular techniques. The total of 151 identified yeasts belonged to 4 different genera: Debaryomyces, Candida, Cryptococcus and Rhodotorula. Debaryomyces hansenii was the most abundant species isolated throughout the whole process as much in the interior as in the exterior of the pieces of three salt levels of "lacón" studied, while Candida zeylanoides was only isolated from the interior of MS and HS batches and from the exterior of LS and HS groups, but at lesser proportion than D. hansenii. Copyright © 2012. Published by Elsevier Ltd.

[Evaluation of bond strength between low fusing porcelain with goldplated cobalt-chromium alloys].

PubMed

Guo, Jing; Zhu, Jia; Zhu, Hong-shui

2014-02-01

To evaluate the bond strength of Vita OMEGA 900 low fusing porcelain fused with the goldplated Wirobond cobalt-chrome metalt ceramic alloy. Low fusing porcelain was fused with the cobalt-chromium alloy strips(group A) and the goldplated cobalt-chromium alloy strips(group B) respectively according to ISO9693 (A:8,B:10). 8 specimens of each group were submitted to three point bending test. Two more test pieces fused with gold plated cobalt-chromium alloys were made (group B'). One test piece of both group B and group B' were observed under scanning electron microscope (SEM) randomly. The data was analyzed with SPSS 16.0 software package. The bond strength (MPa) of group A and group B was 29.92±4.28 and 28.20±5.21, respectively (P>0.05), both higher than 25 MPa required by ISO9693. SEM showed that Vita OMEGA 900 low fusing porcelain and the goldplated Wirobond cobalt-chrome metalt ceramic alloy combined together closely without cracks. Much gold was fused to the cobalt-chrome alloy surface of breaking porcelain specimen after testing. Vita OMEGA 900 low fusing porcelain can match with the goldplated Wirobond cobalt-chrome metalt ceramic alloy. Supported by Foundation of Education Department of Jiangxi Province (GJJ10367).

Distribution of dimorphic yeast species in commercial extra virgin olive oil.

PubMed

Zullo, B A; Cioccia, G; Ciafardini, G

2010-12-01

Recent microbiological research has demonstrated the presence of a rich microflora mainly composed of yeasts in the suspended fraction of freshly produced olive oil. Some of the yeasts are considered useful as they improve the organoleptic characteristics of the oil during preservation, whereas others are considered harmful as they can damage the quality of the oil through the hydrolysis of the triglycerides. However, some dimorphic species can also be found among the unwanted yeasts present in the oil, considered to be opportunistic pathogens to man as they have often been isolated from immunocompromised hospital patients. Present research demonstrates the presence of dimorphic yeast forms in 26% of the commercial extra virgin olive oil originating from different geographical areas, where the dimorphic yeasts are represented by 3-99.5% of the total yeasts. The classified isolates belonged to the opportunistic pathogen species Candida parapsilosis and Candida guilliermondii, while among the dimorphic yeasts considered not pathogenic to man, the Candida diddensiae species was highlighted for the first time in olive oil. The majority of the studied yeast strains resulted lipase positive, and can consequently negatively influence the oil quality through the hydrolysis of the triglycerides. Furthermore, all the strains showed a high level of affinity with some organic solvents and a differing production of biofilm in "vitro" corresponded to a greater or lesser hydrophobia of their cells. Laboratory trials indicated that the dimorphic yeasts studied are sensitive towards some components of the oil among which oleic acid, linoleic acid and triolein, whereas a less inhibiting effect was observed with tricaprilin or when the total polyphenols extracted from the oil were used. The observations carried out on a scanning electron microscope (SEM), demonstrated the production of long un-branched pseudohyphae in all the tested dimorphic yeasts when cultivated on nutrient-deficient substrates. Copyright © 2010 Elsevier Ltd. All rights reserved.

Identification of uncommon oral yeasts from cancer patients by MALDI-TOF mass spectrometry.

PubMed

Aslani, Narges; Janbabaei, Ghasem; Abastabar, Mahdi; Meis, Jacques F; Babaeian, Mahasti; Khodavaisy, Sadegh; Boekhout, Teun; Badali, Hamid

2018-01-08

Opportunistic infections due to Candida species occur frequently in cancer patients because of their inherent immunosuppression. The aim of the present study was to investigate the epidemiology of yeast species from the oral cavity of patients during treatment for oncological and haematological malignancies. MALDI-TOF was performed to identify yeasts isolated from the oral cavity of 350 cancer patients. Moreover, antifungal susceptibility testing was performed in according to CLSI guidelines (M27-A3). Among 162 yeasts and yeast-like fungi isolated from the oral cavity of cancer patients, Candida albicans was the most common species (50.6%), followed by Candida glabrata (24.7%), Pichia kudriavzevii (Candida krusei (9.9%)), Candida tropicalis (4.3%), Candida dubliniensis (3.7%), Kluyveromyces marxianus (Candida kefyr (3.7%)) and Candida parapsilosis (1%). In addition, uncommon yeast species i.e., Saprochaete capitata, Saccharomyces cerevisiae, Clavispora lusitaniae (C. lusitaniae) and Pichia kluyveri (C. eremophila) were recovered from oral lesions. Oral colonization by C. albicans, non-albicans Candida species and uncommon yeasts were as follow; 55%, 44% and 1%, whereas oral infection due to C. albicans was 33.3%, non-albicans Candida species 60.6%, and uncommon yeasts 6.1%. Poor oral hygiene and xerostomia were identified as independent risk factors associated with oral yeast colonization. The overall resistance to fluconazole was 11.7% (19/162). Low MIC values were observed for anidulafungin for all Candida and uncommon yeast species. This current study provides insight into the prevalence and susceptibility profiles of Candida species, including emerging Candida species and uncommon yeasts, isolated from the oral cavity of Iranian cancer patients. The incidence of oral candidiasis was higher amongst patients with hematological malignancies. The majority of oral infections were caused by non-albicans Candida species which were often more resistant to anti-fungal agents. Our findings suggest that anidulafungin should be used as antifungal of choice for prophylaxis in clinically high-risk patients with documented oral colonization or infection.

Comparison of the Petrifilm dry rehydratable film and conventional culture methods for enumeration of yeasts and molds in foods: collaborative study.

PubMed

Knight, M T; Newman, M C; Benzinger, M J; Neufang, K L; Agin, J R; McAllister, J S; Ramos, M

1997-01-01

A collaborative study was performed involving 18 laboratories and 6 food types to compare 3M Petrifilm yeast and mold count plates with the method described in the U.S. Food and Drug Administration's Bacteriological Analytical Manual. Four species of mold and 2 species of yeast were used to inoculate the following foods: hot dogs, corn meal, ketchup, orange juice, yogurt, and cake mix. Each collaborator received 15 samples of each food type: 5 low-level inoculations, 5 high-level inoculations, and 5 uninoculated samples. There was no significant difference between the means of the 2 methods for any product or inoculation level. The Petrifilm yeast and mold count plate method for enumeration of yeasts and molds in foods has been adopted first action by AOAC INTERNATIONAL.

Evolution of the Structure of Local Regions of Fused Metal in Explosion-Welded Nickel-Aluminum Composites Under Heat Treatment

NASA Astrophysics Data System (ADS)

Shmorgun, V. G.; Bogdanov, A. I.; Gurevich, L. M.

2016-03-01

The methods of electron, optical, and atomic force microscopy are used to study the structure, morphology and phase composition of local regions of fused metal in an explosion-welded nickel-aluminum composite. It is shown that the diffusion zone formed due to the heat treatment repeats the contour of the fuse in the first stage and then "absorbs" it upon duration of the hold thus leveling the phase composition. ANi2Al3 Aluminide layer forms on the side of nickel and a NiAl3 layer forms on the side of aluminum.

KSC-99pp0718

NASA Image and Video Library

1999-06-19

At Launch Pad 17A, Cape Canaveral Air Station, workers oversee the lifting of the fairing (right) into the tower. At left is NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite around which the fairing will be fitted. The satellite is scheduled for launch June 24 aboard a Boeing Delta II rocket. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe

KSC-99pp0716

NASA Image and Video Library

1999-06-19

A camera is shown mounted on the second stage of the Boeing Delta II rocket scheduled to launch NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite June 24 from Launch Pad 17A, Cape Canaveral Air Station. The camera will record the separation of the fairing encircling the satellite, which should occur several minutes after launch. FUSE is designed to scour the cosmos for the fossil record of the origins of the universe hydrogen and deuterium. Scientists will use FUSE to study hydrogen and deuterium to unlock the secrets of how the primordial chemical elements of which all stars, planets and life evolved, were created and distributed since the birth of the universe

Risk assessment for the spread of Candida sp. in dental chair unit waterlines using molecular techniques.

PubMed

Mazari, Wissame; Boucherit-Otmani, Zahia; El Haci, Imad Abdelhamid; Ilahi, Amine; Boucherit, Kebir

2018-05-04

The aim of this study was to evaluate the presence of yeasts in dental chair unit waterlines (DCUWLs) and to test their ability to form biofilms. Eighteen dental waterlines were analysed by culture in liquid Sabouraud in order to allow the quantification and the purification of isolated yeasts from their internal surfaces. All isolates were identified by standard laboratory procedures, including CHROMagar Candida medium for orientation, commercial yeast identification system Api Candida, MALDI-TOF MS and DNA sequencing. To evaluate their kinetics of antifungal susceptibility during different phases of biofilm formation, these yeasts were subjected to three antifungal agents. From the 18 DCUWLs studied, 10 were altered (55.56%). Eleven strains of Candida sp. [Candida albicans (2), Candida guilliermondii (5) and Candida glabrata (4)] and two species of non-Candida; Rhodotorula spp. (1) and Trichosporon spp. (2) were identified. The majority of yeasts in planktonic form were susceptible to amphotericin B, caspofungin and voriconazole, except C. albicans was resistant to voriconazole. In the biofilm form, caspofungin was the most effective antifungal agent for all isolated strains. For the other antifungal agents, sessile cells were resistant. Several types of yeasts were identified; the most frequently isolated genus was Candida. The majority of these yeasts had the ability to form biofilms and resisted antifungal agents used in this study. © 2018 FDI World Dental Federation.

Use of Non-Conventional Cell Disruption Method for Extraction of Proteins from Black Yeasts

PubMed Central

Čolnik, Maja; Primožič, Mateja; Knez, Željko; Leitgeb, Maja

2016-01-01

The influence of pressure and treatment time on cells disruption of different black yeasts and on activities of extracted proteins using supercritical carbon dioxide process was studied. The cells of three different black yeasts Phaeotheca triangularis, Trimatostroma salinum, and Wallemia ichthyophaga were exposed to supercritical carbon dioxide (SC CO2) by varying pressure at fixed temperature (35°C). The black yeasts cell walls were disrupted, and the content of the cells was spilled into the liquid medium. The impact of SC CO2 conditions on secretion of enzymes and proteins from black yeast cells suspension was studied. The residual activity of the enzymes cellulase, β-glucosidase, α-amylase, and protease was studied by enzymatic assay. The viability of black yeast cells was determined by measuring the optical density of the cell suspension at 600 nm. The total protein concentration in the suspension was determined on UV–Vis spectrophotometer at 595 nm. The release of intracellular and extracellular products from black yeast cells was achieved. Also, the observation by an environmental scanning electron microscopy shows major morphological changes with SC CO2-treated cells. The advantages of the proposed method are in a simple use, which is also possible for heat-sensitive materials on one hand and on the other hand integration of the extraction of enzymes and their use in biocatalytical reactions. PMID:27148527

Schwanniomyces Klocker emend. M. Suzuki & Kurtzman (2010)

USDA-ARS?s Scientific Manuscript database

This chapter describes the ascomycete yeast genus Schwanniomyces and is to be published in The Yeasts, A Taxonomic Study, 5th edn. The genus Schwanniomyces has seven assigned species, many worldwide in distribution. Schwanniomyces is one of the few yeast genera with species that seem to live in th...

Combining mutualistic yeast and pathogenic virus - a novel method for control for codling moth control

USDA-ARS?s Scientific Manuscript database

Studies evaluated the lethal effectiveness of combining yeasts isolated from larvae of codling moth, Cydia pomonella (L.) with the codling moth granulosis virus (CpGV). Apples were treated with CpGV and three yeast species, including Metschnikowia pulcherrima Pitt and Miller, Cryptococcus tephrensis...

Yeast supplements enhance immune function and performance in food-producing livestock: A review

USDA-ARS?s Scientific Manuscript database

More livestock producers are seeking natural alternatives to antibiotics and antimicrobials, and searching for supplements to enhance growth performance, and overall animal health and well-being. Some of the compounds currently being utilized and studied are live yeast and yeast-based products deriv...

Accumulation and metabolism of selenium by yeast cells.

PubMed

Kieliszek, Marek; Błażejak, Stanisław; Gientka, Iwona; Bzducha-Wróbel, Anna

2015-07-01

This paper examines the process of selenium bioaccumulation and selenium metabolism in yeast cells. Yeast cells can bind elements in ionic from the environment and permanently integrate them into their cellular structure. Up to now, Saccharomyces cerevisiae, Candida utilis, and Yarrowia lipolytica yeasts have been used primarily in biotechnological studies to evaluate binding of minerals. Yeast cells are able to bind selenium in the form of both organic and inorganic compounds. The process of bioaccumulation of selenium by microorganisms occurs through two mechanisms: extracellular binding by ligands of membrane assembly and intracellular accumulation associated with the transport of ions across the cytoplasmic membrane into the cell interior. During intracellular metabolism of selenium, oxidation, reduction, methylation, and selenoprotein synthesis processes are involved, as exemplified by detoxification processes that allow yeasts to survive under culture conditions involving the elevated selenium concentrations which were observed. Selenium yeasts represent probably the best absorbed form of this element. In turn, in terms of wide application, the inclusion of yeast with accumulated selenium may aid in lessening selenium deficiency in a diet.

Analysis of the yeast short-term Crabtree effect and its origin

PubMed Central

Hagman, Arne; Säll, Torbjörn; Piškur, Jure

2014-01-01

The short-term Crabtree effect is defined as the immediate occurrence of aerobic alcoholic fermentation in response to provision of a pulse of excess sugar to sugar-limited yeast cultures. Here we have characterized ten yeast species with a clearly defined phylogenetic relationship. Yeast species were cultivated under glucose-limited conditions, and we studied their general carbon metabolism in response to a glucose pulse. We generated an extensive collection of data on glucose and oxygen consumption, and ethanol and carbon dioxide generation. We conclude that the Pichia,Debaryomyces,Eremothecium and Kluyveromyces marxianus yeasts do not exhibit any significant ethanol formation, while Kluyveromyces lactis behaves as an intermediate yeast, and Lachancea,Torulaspora,Vanderwaltozyma and Saccharomyces yeasts exhibit rapid ethanol accumulation. Based on the present data and our previous data relating to the presence of the long-term Crabtree effect in over 40 yeast species, we speculate that the origin of the short-term effect may coincide with the origin of the long-term Crabtree effect in the Saccharomycetales lineage, occurring ∼ 150 million years ago. PMID:25161062

Culturable yeasts in meltwaters draining from two glaciers in the Italian Alps

NASA Astrophysics Data System (ADS)

Buzzini, Pietro; Turchetti, Benedetta; Diolaiuti, Guglielmina; D'Agata, Carlo; Martini, Alessandro; Smiraglia, Claudio

The meltwaters draining from two glaciers in the Italian Alps contain metabolically active yeasts isolable by culture-based laboratory procedures. The average number of culturable yeast cells in the meltwaters was 10 20 colony-forming units (CFU) L-1, whereas supraglacial stream waters originating from overlying glacier ice contained <1 CFU L-1. Yeast cell number increased as the suspended-sediment content of the water samples increased. Basidiomycetous yeasts represent >80% of isolated strains (Cryptococcus spp. and Rhodotorula spp. were 33.3% and 17.8% of total strains, respectively). Culturable yeasts were psychrotolerant, predominantly obligate aerobes and able to degrade organic macromolecules (e.g. starch, esters, lipids, proteins). To the authors' knowledge, this is the first study to report the presence of culturable yeasts in meltwaters originating from glaciers. On the basis of these results, it is reasonable to suppose that the viable yeasts observed in meltwaters derived predominantly from the subglacial zone and that they originated from the subglacial microbial community. Their metabolic abilities could contribute to the microbial activity occurring in subglacial environments.

Feasibility of protein turnover studies in prototroph Saccharomyces cerevisiae strains.

PubMed

Martin-Perez, Miguel; Villén, Judit

2015-04-07

Quantitative proteomics studies of yeast that use metabolic labeling with amino acids rely on auxotrophic mutations of one or more genes on the amino acid biosynthesis pathways. These mutations affect yeast metabolism and preclude the study of some biological processes. Overcoming this limitation, it has recently been described that proteins in a yeast prototrophic strain can also be metabolically labeled with heavy amino acids. However, the temporal profiles of label incorporation under the different phases of the prototroph's growth have not been examined. Labeling trajectories are important in the study of protein turnover and dynamics, in which label incorporation into proteins is monitored across many time points. Here we monitored protein labeling trajectories for 48 h after a pulse with heavy lysine in a yeast prototrophic strain and compared them with those of a lysine auxotrophic yeast. Labeling was successful in prototroph yeast during exponential growth phase but not in stationary phase. Furthermore, we were able to determine the half-lives of more than 1700 proteins during exponential phase of growth with high accuracy and reproducibility. We found a median half-life of 2 h in both strains, which corresponds with the cellular doubling time. Nucleolar and ribosomal proteins showed short half-lives, whereas mitochondrial proteins and other energy production enzymes presented longer half-lives. Except for some proteins involved in lysine biosynthesis, we observed a high correlation in protein half-lives between prototroph and auxotroph strains. Overall, our results demonstrate the feasibility of using prototrophs for proteomic turnover studies and provide a reliable data set of protein half-lives in exponentially growing yeast.

Effect of chromium supplementation on glycated hemoglobin and fasting plasma glucose in patients with diabetes mellitus.

PubMed

Yin, Raynold V; Phung, Olivia J

2015-02-13

Chromium (Cr) is a trace element involved in glucose homeostasis. We aim to evaluate and quantify the effects of Cr supplementation on A1C and FPG in patients with T2DM. A systematic literature search of Pubmed, EMBASE and the Cochrane Library (from database inception to 11/2014) with no language restrictions sought RCTs or cohort studies evaluating Cr supplementation in T2DM vs control and reporting either change in glycated hemoglobin (A1C) or fasting plasma glucose (FPG). Meta-analysis was conducted on each subtype of Cr supplement separately, and was analyzed by random effects model to yield the weighted mean differences (WMD) and 95% confidence intervals (CIs). Heterogeneity was assessed by using the I(2) statistic. A total of 14 RCTs (n=875 participants, mean age range: 30 to 83 years old, 8 to 24 weeks of follow-up) were identified (Cr chloride: n=3 study, Cr picolinate: n=5 study, brewer's yeast: n=4 study and Cr yeast: n=3 study). Compared with placebo, Cr yeast, brewer's yeast and Cr picolinate did not show statistically significant effects on A1C. Furthermore, compared to control, Cr chloride, Cr yeast and Cr picolinate showed no effect on FPG, however, brewer's yeast showed a statistically significant decrease in FPG -19.23 mg/dL (95% CI=-35.30 to -3.16, I(2)=21%, n=137). Cr supplementation with brewer's yeast may provide marginal benefits in lowering FPG in patients with T2DM compared to placebo however it did not have any effect on A1C.

Studies of the expression of human poly(ADP-ribose) polymerase-1 in Saccharomyces cerevisiae and identification of PARP-1 substrates by yeast proteome microarray screening.

PubMed

Tao, Zhihua; Gao, Peng; Liu, Hung-Wen

2009-12-15

Poly(ADP-ribosyl)ation of various nuclear proteins catalyzed by a family of NAD(+)-dependent enzymes, poly(ADP-ribose) polymerases (PARPs), is an important posttranslational modification reaction. PARP activity has been demonstrated in all types of eukaryotic cells with the exception of yeast, in which the expression of human PARP-1 was shown to lead to retarded cell growth. We investigated the yeast growth inhibition caused by human PARP-1 expression in Saccharomyces cerevisiae. Flow cytometry analysis reveals that PARP-1-expressing yeast cells accumulate in the G(2)/M stage of the cell cycle. Confocal microscopy analysis shows that human PARP-1 is distributed throughout the nucleus of yeast cells but is enriched in the nucleolus. Utilizing yeast proteome microarray screening, we identified 33 putative PARP-1 substrates, six of which are known to be involved in ribosome biogenesis. The poly(ADP-ribosyl)ation of three of these yeast proteins, together with two human homologues, was confirmed by an in vitro PARP-1 assay. Finally, a polysome profile analysis using sucrose gradient ultracentrifugation demonstrated that the ribosome levels in yeast cells expressing PARP-1 are lower than those in control yeast cells. Overall, our data suggest that human PARP-1 may affect ribosome biogenesis by modifying certain nucleolar proteins in yeast. The artificial PARP-1 pathway in yeast may be used as a simple platform to identify substrates and verify function of this important enzyme.

Microbial Interactions within a Cheese Microbial Community▿ †

PubMed Central

Mounier, Jérôme; Monnet, Christophe; Vallaeys, Tatiana; Arditi, Roger; Sarthou, Anne-Sophie; Hélias, Arnaud; Irlinger, Françoise

2008-01-01

The interactions that occur during the ripening of smear cheeses are not well understood. Yeast-yeast interactions and yeast-bacterium interactions were investigated within a microbial community composed of three yeasts and six bacteria found in cheese. The growth dynamics of this community was precisely described during the ripening of a model cheese, and the Lotka-Volterra model was used to evaluate species interactions. Subsequently, the effects on ecosystem functioning of yeast omissions in the microbial community were evaluated. It was found both in the Lotka-Volterra model and in the omission study that negative interactions occurred between yeasts. Yarrowia lipolytica inhibited mycelial expansion of Geotrichum candidum, whereas Y. lipolytica and G. candidum inhibited Debaryomyces hansenii cell viability during the stationary phase. However, the mechanisms involved in these interactions remain unclear. It was also shown that yeast-bacterium interactions played a significant role in the establishment of this multispecies ecosystem on the cheese surface. Yeasts were key species in bacterial development, but their influences on the bacteria differed. It appeared that the growth of Arthrobacter arilaitensis or Hafnia alvei relied less on a specific yeast function because these species dominated the bacterial flora, regardless of which yeasts were present in the ecosystem. For other bacteria, such as Leucobacter sp. or Brevibacterium aurantiacum, growth relied on a specific yeast, i.e., G. candidum. Furthermore, B. aurantiacum, Corynebacterium casei, and Staphylococcus xylosus showed reduced colonization capacities in comparison with the other bacteria in this model cheese. Bacterium-bacterium interactions could not be clearly identified. PMID:17981942

Isolation and Identification of the Indigenous Yeast Population during Spontaneous Fermentation of Isabella (Vitis labrusca L.) Grape Must.

PubMed

Raymond Eder, María L; Reynoso, Cristina; Lauret, Santiago C; Rosa, Alberto L

2017-01-01

Grape must harbors a complex community of yeast species responsible for spontaneous alcoholic fermentation. Although there are detailed studies on the microbiota of Vitis vinifera L. grapes, less is known about the diversity and behavior of yeast communities present on fermenting grape must from other species of Vitis . In this work, we used a culture-dependent method to study the identity and dynamics of the indigenous yeast population present during the spontaneous fermentation of Isabella ( Vitis labrusca L.) grape must. Alcoholic fermentation was conducted using standard enological practices, and the associated non- Saccharomyces and S. cerevisiae yeast community was analyzed using selective growth media and 5.8-ITS DNA sequencing. Candida californica, Candida hellenica, Starmerella bacillaris (synonym Candida zemplinina ), Hanseniaspora uvarum , and Hanseniaspora vineae were the main non- Saccharomyces species identified on Isabella fermenting must. Issatchenkia hanoiensis , a yeast species rarely found on Vitis vinifera L. grapes, was also recognized on Isabella grape must. Candida azymoides, Candida californica and Pichia cecembensis , identified in this work on Isabella fermenting must, have not previously been found on Vitis vinifera L. grape must. Interestingly, C. azymoides, I. hanoiensis and P. cecembensis have recently been isolated from the surface of Vitis labrusca L. grapes from vineyards in the Azores archipelago, suggesting that specific Vitis -yeast species associations are formed independently of geographic origin. We suggest that C. azymoides, C. californica , and P. cecembensis are yeast species preferentially associated with Vitis labrusca L. grapes. Specific biological interactions between grapevines and yeast species may underlie the assembly of differential Vitis -microbial communities.

Isolation and Identification of the Indigenous Yeast Population during Spontaneous Fermentation of Isabella (Vitis labrusca L.) Grape Must

PubMed Central

Raymond Eder, María L.; Reynoso, Cristina; Lauret, Santiago C.; Rosa, Alberto L.

2017-01-01

Grape must harbors a complex community of yeast species responsible for spontaneous alcoholic fermentation. Although there are detailed studies on the microbiota of Vitis vinifera L. grapes, less is known about the diversity and behavior of yeast communities present on fermenting grape must from other species of Vitis. In this work, we used a culture-dependent method to study the identity and dynamics of the indigenous yeast population present during the spontaneous fermentation of Isabella (Vitis labrusca L.) grape must. Alcoholic fermentation was conducted using standard enological practices, and the associated non-Saccharomyces and S. cerevisiae yeast community was analyzed using selective growth media and 5.8-ITS DNA sequencing. Candida californica, Candida hellenica, Starmerella bacillaris (synonym Candida zemplinina), Hanseniaspora uvarum, and Hanseniaspora vineae were the main non-Saccharomyces species identified on Isabella fermenting must. Issatchenkia hanoiensis, a yeast species rarely found on Vitis vinifera L. grapes, was also recognized on Isabella grape must. Candida azymoides, Candida californica and Pichia cecembensis, identified in this work on Isabella fermenting must, have not previously been found on Vitis vinifera L. grape must. Interestingly, C. azymoides, I. hanoiensis and P. cecembensis have recently been isolated from the surface of Vitis labrusca L. grapes from vineyards in the Azores archipelago, suggesting that specific Vitis-yeast species associations are formed independently of geographic origin. We suggest that C. azymoides, C. californica, and P. cecembensis are yeast species preferentially associated with Vitis labrusca L. grapes. Specific biological interactions between grapevines and yeast species may underlie the assembly of differential Vitis-microbial communities. PMID:28424672

Genomic signatures of adaptation to wine biological ageing conditions in biofilm-forming flor yeasts.

PubMed

Coi, A L; Bigey, F; Mallet, S; Marsit, S; Zara, G; Gladieux, P; Galeote, V; Budroni, M; Dequin, S; Legras, J L

2017-04-01

The molecular and evolutionary processes underlying fungal domestication remain largely unknown despite the importance of fungi to bioindustry and for comparative adaptation genomics in eukaryotes. Wine fermentation and biological ageing are performed by strains of S. cerevisiae with, respectively, pelagic fermentative growth on glucose and biofilm aerobic growth utilizing ethanol. Here, we use environmental samples of wine and flor yeasts to investigate the genomic basis of yeast adaptation to contrasted anthropogenic environments. Phylogenetic inference and population structure analysis based on single nucleotide polymorphisms revealed a group of flor yeasts separated from wine yeasts. A combination of methods revealed several highly differentiated regions between wine and flor yeasts, and analyses using codon-substitution models for detecting molecular adaptation identified sites under positive selection in the high-affinity transporter gene ZRT1. The cross-population composite likelihood ratio revealed selective sweeps at three regions, including in the hexose transporter gene HXT7, the yapsin gene YPS6 and the membrane protein coding gene MTS27. Our analyses also revealed that the biological ageing environment has led to the accumulation of numerous mutations in proteins from several networks, including Flo11 regulation and divalent metal transport. Together, our findings suggest that the tuning of FLO11 expression and zinc transport networks are a distinctive feature of the genetic changes underlying the domestication of flor yeasts. Our study highlights the multiplicity of genomic changes underlying yeast adaptation to man-made habitats and reveals that flor/wine yeast lineage can serve as a useful model for studying the genomics of adaptive divergence. © 2017 John Wiley & Sons Ltd.

Study the effect of Vitamin K on intracellular NAD level in yeast by fluorescence spectrum

NASA Astrophysics Data System (ADS)

Yahong, Chen; Ruxiu, Cai; Ke, Zhang

2007-05-01

The intracellular NAD level plays a pivotal role in numerous biological processes such as rhythm, senescence, cancer and death. The study of the intracellular NAD level has been one of the "hotspots" in biomedical research. We investigated the effect of Vitamin K on intracellular NAD level in yeast by fluorescence spectrum in this paper. Plasma membrane redox system of yeast was found to be greatly promoted by the addition of Vitamin K 3 or Vitamin K 1. Ferricyanide reduction catalyzed by Vitamin K was accompanied by the decrease in intracellular NADH concentration and the increase in intracellular NAD level of yeast cells.

Screening and evaluation of the glucoside hydrolase activity in Saccharomyces and Brettanomyces brewing yeasts.

PubMed

Daenen, L; Saison, D; Sterckx, F; Delvaux, F R; Verachtert, H; Derdelinckx, G

2008-02-01

The aim of this study was to select and examine Saccharomyces and Brettanomyces brewing yeasts for hydrolase activity towards glycosidically bound volatile compounds. A screening for glucoside hydrolase activity of 58 brewing yeasts belonging to the genera Saccharomyces and Brettanomyces was performed. The studied Saccharomyces brewing yeasts did not show 1,4-beta-glucosidase activity, but a strain dependent beta-glucanase activity was observed. Some Brettanomyces species did show 1,4-beta-glucosidase activity. The highest constitutive activity was found in Brettanomyces custersii. For the most interesting strains the substrate specificity was studied and their activity was evaluated in fermentation experiments with added hop glycosides. Fermentations with Br. custersii led to the highest release of aglycones. Pronounced exo-beta-glucanase activity in Saccharomyces brewing yeasts leads to a higher release of certain aglycones. Certain Brettanomyces brewing yeasts, however, are more interesting for hydrolysis of glycosidically bound volatiles of hops. The release of flavour active compounds from hop glycosides opens perspectives for the bioflavouring and product diversification of beverages like beer. The release can be enhanced by using Saccharomyces strains with high exo-beta-glucanase activity. Higher activities can be found in Brettanomyces species with beta-glucosidase activity.

A long term field study of the effect of fungicides penconazole and sulfur on yeasts in the vineyard.

PubMed

Cordero-Bueso, Gustavo; Arroyo, Teresa; Valero, Eva

2014-10-17

This research deals with how two fungicide treatments against powdery mildew, penconazole as a systematic fungicide and sulfur as an inorganic broad-spectrum fungicide, affect the diversity and density of wine yeasts associated with grape berry surfaces and subsequent spontaneous fermentations. Unlike other studies in this area, this work aims to evaluate this effect on the population dynamics in the environment, the conditions of which are not reproducible in the laboratory. A long term (three year) sampling plan was thus devised. A minimum inhibitory concentration assay was also carried out in the laboratory in order to prove the influence of these antifungals on yeast populations. While both antifungal treatments (penconazole and sulfur) were similarly effective against powdery mildew, each had a very different effect on yeast populations. Penconazole showed the most negative effect on biodiversity in the vineyard and was the fungicide to which the isolated yeasts showed the greatest sensitivity. This study therefore evidences the suitability of treatment with sulfur, in both conventional and organic viticulture, to preserve the yeast population associated with grape berries, in particular the Saccharomyces cerevisiae species. Copyright © 2014 Elsevier B.V. All rights reserved.

31 CFR 100.12 - Exchange of fused and mixed coins.

Code of Federal Regulations, 2014 CFR

2014-07-01

... 31 Money and Finance: Treasury 1 2014-07-01 2014-07-01 false Exchange of fused and mixed coins... of fused and mixed coins. (a) Definitions. (1) Fused coins are U.S. coins which are melted to the....S. coins. (2) Mixed coins are U.S. coins of several alloy categories which are presented together...

31 CFR 100.12 - Exchange of fused and mixed coins.

Code of Federal Regulations, 2011 CFR

2011-07-01

... 31 Money and Finance: Treasury 1 2011-07-01 2011-07-01 false Exchange of fused and mixed coins... of fused and mixed coins. (a) Definitions. (1) Fused coins are U.S. coins which are melted to the....S. coins. (2) Mixed coins are U.S. coins of several alloy categories which are presented together...

31 CFR 100.12 - Exchange of fused and mixed coins.

Code of Federal Regulations, 2012 CFR

2012-07-01

... 31 Money and Finance: Treasury 1 2012-07-01 2012-07-01 false Exchange of fused and mixed coins... of fused and mixed coins. (a) Definitions. (1) Fused coins are U.S. coins which are melted to the....S. coins. (2) Mixed coins are U.S. coins of several alloy categories which are presented together...

31 CFR 100.12 - Exchange of fused and mixed coins.

Code of Federal Regulations, 2013 CFR

2013-07-01

... 31 Money and Finance: Treasury 1 2013-07-01 2013-07-01 false Exchange of fused and mixed coins... of fused and mixed coins. (a) Definitions. (1) Fused coins are U.S. coins which are melted to the....S. coins. (2) Mixed coins are U.S. coins of several alloy categories which are presented together...

Development of Fuses for Protection of Geiger-Mode Avalanche Photodiode Arrays

NASA Astrophysics Data System (ADS)

Grzesik, Michael; Bailey, Robert; Mahan, Joe; Ampe, Jim

2015-11-01

Current-limiting fuses composed of Ti/Al/Ni were developed for use in Geiger-mode avalanche photodiode arrays for each individual pixel in the array. The fuses were designed to burn out at ˜4.5 × 10-3 A and maintain post-burnout leakage currents less than 10-7 A at 70 V sustained for several minutes. Experimental fuse data are presented and successful incorporation of the fuses into a 256 × 64 pixel InP-based Geiger-mode avalanche photodiode array is reported.

Integrated fuses for OLED lighting device

DOEpatents

Pschenitzka, Florian [San Jose, CA

2007-07-10

An embodiment of the present invention pertains to an electroluminescent lighting device for area illumination. The lighting device is fault tolerant due, in part, to the patterning of one or both of the electrodes into strips, and each of one or more of these strips has a fuse formed on it. The fuses are integrated on the substrate. By using the integrated fuses, the number of external contacts that are used is minimized. The fuse material is deposited using one of the deposition techniques that is used to deposit the thin layers of the electroluminescent lighting device.

Population analysis of biofilm yeasts during fino sherry wine aging in the Montilla-Moriles D.O. region.

PubMed

Marin-Menguiano, Miriam; Romero-Sanchez, Sandra; Barrales, Ramón R; Ibeas, Jose I

2017-03-06

Fino is the most popular sherry wine produced in southern Spain. Fino is matured by biological aging under a yeast biofilm constituted of Saccharomyces cerevisiae yeasts. Although different S. cerevisiae strains can be identified in such biofilms, their diversity and contribution to wine character have been poorly studied. In this work, we analyse the flor yeast population in five different wineries from the Montilla-Moriles D.O. (Denominación de Origen) in southern Spain. Yeasts present in wines of different ages were identified using two different culture-dependent molecular techniques. From 2000 individual yeast isolates, five different strains were identified with one of them dominating in four out of the five wineries analysed, and representing 76% of all the yeast isolates collected. Surprisingly, this strain is similar to the predominant strain isolated twenty years ago in Jerez D.O. wines, suggesting that this yeast is particularly able to adapt to such a stressful environment. Fino wine produced with pure cultures of three of the isolated strains resulted in different levels of acetaldehyde. Because acetaldehyde levels are a distinctive characteristic of fino wines and an indicator of fino aging, the use of molecular techniques for yeast identification and management of yeast populations may be of interest for fino wine producers looking to control one of the main features of this wine. Copyright © 2016 Elsevier B.V. All rights reserved.

Harvesting yeast (Saccharomyces cerevisiae) at different physiological phases significantly affects its functionality in bread dough fermentation.

PubMed

Rezaei, Mohammad N; Dornez, Emmie; Jacobs, Pieter; Parsi, Anali; Verstrepen, Kevin J; Courtin, Christophe M

2014-05-01

Fermentation of sugars into CO2, ethanol and secondary metabolites by baker's yeast (Saccharomyces cerevisiae) during bread making leads to leavening of dough and changes in dough rheology. The aim of this study was to increase our understanding of the impact of yeast on dough related aspects by investigating the effect of harvesting yeast at seven different points of the growth profile on its fermentation performance, metabolite production, and the effect on critical dough fermentation parameters, such as gas retention potential. The yeast cells harvested during the diauxic shift and post-diauxic growth phase showed a higher fermentation rate and, consequently, higher maximum dough height than yeast cells harvested in the exponential or stationary growth phase. The results further demonstrate that the onset of CO2 loss from fermenting dough is correlated with the fermentation rate of yeast, but not with the amount of CO2 that accumulated up to the onset point. Analysis of the yeast metabolites produced in dough yielded a possible explanation for this observation, as they are produced in different levels depending on physiological phase and in concentrations that can influence dough matrix properties. Together, our results demonstrate a strong effect of yeast physiology at the time of harvest on subsequent dough fermentation performance, and hint at an important role of yeast metabolites on the subsequent gas holding capacity. Copyright © 2013 Elsevier Ltd. All rights reserved.

Water quality and antifungal susceptibility of opportunistic yeast pathogens from rivers.

PubMed

Monapathi, M E; Bezuidenhout, C C; Rhode, O H J

2017-03-01

Yeasts from water sources have been associated with diseases ranging from superficial mucosal infections to life threatening diseases. The aim of this study was to determine the water quality as well as diversity and antifungal susceptibility of yeasts from two rivers. Yeast levels and physico-chemical parameter data were analyzed by principal component analysis to determine correlations between physico-chemical data and yeast levels. Yeast morphotypes were identified by biochemical tests and 26S rRNA gene sequencing. Disk diffusion antifungal susceptibility tests were conducted. Physico-chemical parameters of the water were within target water quality range (TWQR) for livestock farming. For irrigational use, total dissolved solids and nitrates were not within the TWQR. Yeast levels ranged between 27 ± 10 and 2,573 ± 306 cfu/L. Only non-pigmented, ascomycetous yeasts were isolated. Saccharomyces cerevisiae and Candida glabrata were most frequently isolated. Several other opportunistic pathogens were also isolated. A large number of isolates were resistant to azoles, especially fluconazole, but also to other antifungal classes. Candida species were resistant to almost all the antifungal classes. These water sources are used for recreation and religious as well as for watering livestock and irrigation. Of particular concern is the direct contact of individuals with opportunistic yeast, especially the immune-compromised. Resistance of these yeast species to antifungal agents is a further health concern.

Homo- and heterofermentative lactobacilli differently affect sugarcane-based fuel ethanol fermentation.

PubMed

Basso, Thiago Olitta; Gomes, Fernanda Sgarbosa; Lopes, Mario Lucio; de Amorim, Henrique Vianna; Eggleston, Gillian; Basso, Luiz Carlos

2014-01-01

Bacterial contamination during industrial yeast fermentation has serious economic consequences for fuel ethanol producers. In addition to deviating carbon away from ethanol formation, bacterial cells and their metabolites often have a detrimental effect on yeast fermentative performance. The bacterial contaminants are commonly lactic acid bacteria (LAB), comprising both homo- and heterofermentative strains. We have studied the effects of these two different types of bacteria upon yeast fermentative performance, particularly in connection with sugarcane-based fuel ethanol fermentation process. Homofermentative Lactobacillus plantarum was found to be more detrimental to an industrial yeast strain (Saccharomyces cerevisiae CAT-1), when compared with heterofermentative Lactobacillus fermentum, in terms of reduced yeast viability and ethanol formation, presumably due to the higher titres of lactic acid in the growth medium. These effects were only noticed when bacteria and yeast were inoculated in equal cell numbers. However, when simulating industrial fuel ethanol conditions, as conducted in Brazil where high yeast cell densities and short fermentation time prevail, the heterofermentative strain was more deleterious than the homofermentative type, causing lower ethanol yield and out competing yeast cells during cell recycle. Yeast overproduction of glycerol was noticed only in the presence of the heterofermentative bacterium. Since the heterofermentative bacterium was shown to be more deleterious to yeast cells than the homofermentative strain, we believe our findings could stimulate the search for more strain-specific antimicrobial agents to treat bacterial contaminations during industrial ethanol fermentation.

Nectar yeasts warm the flowers of a winter-blooming plant

PubMed Central

Herrera, Carlos M.; Pozo, María I.

2010-01-01

Yeasts are ubiquitous in terrestrial and aquatic microbiota, yet their ecological functionality remains relatively unexplored in comparison with other micro-organisms. This paper formulates and tests the novel hypothesis that heat produced by the sugar catabolism of yeast populations inhabiting floral nectar can increase the temperature of floral nectar and, more generally, modify the within-flower thermal microenvironment. Two field experiments were designed to test this hypothesis for the winter-blooming herb Helleborus foetidus (Ranunculaceae). In experiment 1, the effect of yeasts on the within-flower thermal environment was tested by excluding them from flowers, while in experiment 2 the test involved artificial inoculation of virgin flowers with yeasts. Nectary temperature (Tnect), within-flower air temperature (Tflow) and external air temperature (Tair) were measured on experimental and control flowers in both experiments. Experimental exclusion of yeasts from the nectaries significantly reduced, and experimental addition of yeasts significantly increased, the temperature excess of nectaries (ΔTnect = Tnect − Tair) and the air space inside flowers in relation to the air just outside the flowers. In non-experimental flowers exposed to natural pollinator visitation, ΔTnect was linearly related to log yeast cell density in nectar, and reached +6°C in nectaries with the densest yeast populations. The warming effect of nectar-dwelling yeasts documented in this study suggests novel ecological mechanisms potentially linking nectarivorous microbes with winter-blooming plants and their insect pollinators. PMID:20147331

New insights on the baker's yeast-mediated hydration of oleic acid: the bacterial contaminants of yeast are responsible for the stereoselective formation of (R)-10-hydroxystearic acid.

PubMed

Serra, S; De Simeis, D

2018-03-01

The preparation of the high-value flavour γ-dodecalactone is based on the biotransformation of natural 10-HSA, which is in turn obtained by microbial hydration of oleic acid. We want to establish a reliable baker's yeast-mediated procedure for 10-HSA preparation. The previously reported yeast-mediated hydration procedures are unreliable because bacteria-free baker's yeast is not able to hydrate oleic acid. The actual responsible for performing this reaction are the bacterial contaminants present in baker's yeast. Moreover, we demonstrated that the enantioselectivity in the production of (R)-10-HSA is affected mainly by the temperature used in the biotransformation. We demonstrated that Saccharomyces cerevisiae is not able to hydrate oleic acid, whereas different bacterial strains present in baker's yeast transform oleic acid into (R)-10-HSA. We reported a general procedure for the preparation of (R)-10-HSA starting from oleic acid and using commercially available baker's yeast. This study holds both scientific and industrial interest. It unambiguously establishes that the eukaryote micro-organisms present in baker's yeast are not able to hydrate oleic acid. The isolation of oleic acid hydrating bacterial strains from commercial baker's yeast points to their prospective use for the industrial synthesis of 10-HSA. © 2017 The Society for Applied Microbiology.

TAT improves in vitro transportation of fortilin through midgut and into hemocytes of white shrimp Litopenaeus vannamei

NASA Astrophysics Data System (ADS)

Zhou, Yi; Zhang, Wenbing; Mai, Kangsen; Xu, Wei; Zhang, Yanjiao; Ai, Qinghui; Wang, Xiaojie

2012-06-01

Fortilin is a multifunctional protein implicated in many important cellular processes. Since injection of Pm-fortilin reduces shrimp mortality caused by white spot syndrome virus (WSSV), there is potential application of fortilin in shrimp culture. In the present study, in order to improve trans-membrane transportation efficiency, the protein transduction domain of the transactivator of transcription (TAT) peptide was fused to fortilin. The Pichia pastoris yeast expression system, which is widely accepted in animal feeds, was used for production of recombinant fusion protein. Green fluorescence protein (GFP) was selected as a reporter because of its intrinsic visible fluorescence. The fortilin, TAT and GFP fusion protein were constructed. Their trans-membrane transportation efficiency and effects on immune response of shrimp were analyzed in vitro. Results showed that TAT peptide improved in vitro uptake of fortilin into the hemocytes and midgut of Litopenaeus vannamei. The phenoloxidase (PO) activity of hemocytes incubated with GFP-Fortilin or GFP-Fortilin-TAT was significantly increased compared with that in the control without expressed fortilin. The PO activity of hemocytes incubated with 200 μg mL-1 GFP-Fortilin-TAT was significantly higher than that in the group with the same concentration of GFP-Fortilin. Hemocytes incubated with GFP-Fortilin-TAT at all concentrations showed significantly higher nitric oxide synthase (NOS) activity than those in the control or in the GFP-Fortilin treatment. The present in vitro study indicated that TAT fusion protein improved the immune effect of fortilin.

Industrial Relevance of Chromosomal Copy Number Variation in Saccharomyces Yeasts.

PubMed

Gorter de Vries, Arthur R; Pronk, Jack T; Daran, Jean-Marc G

2017-06-01

Chromosomal copy number variation (CCNV) plays a key role in evolution and health of eukaryotes. The unicellular yeast Saccharomyces cerevisiae is an important model for studying the generation, physiological impact, and evolutionary significance of CCNV. Fundamental studies of this yeast have contributed to an extensive set of methods for analyzing and introducing CCNV. Moreover, these studies provided insight into the balance between negative and positive impacts of CCNV in evolutionary contexts. A growing body of evidence indicates that CCNV not only frequently occurs in industrial strains of Saccharomyces yeasts but also is a key contributor to the diversity of industrially relevant traits. This notion is further supported by the frequent involvement of CCNV in industrially relevant traits acquired during evolutionary engineering. This review describes recent developments in genome sequencing and genome editing techniques and discusses how these offer opportunities to unravel contributions of CCNV in industrial Saccharomyce s strains as well as to rationally engineer yeast chromosomal copy numbers and karyotypes. Copyright © 2017 Gorter de Vries et al.

Industrial Relevance of Chromosomal Copy Number Variation in Saccharomyces Yeasts

PubMed Central

Gorter de Vries, Arthur R.; Pronk, Jack T.

2017-01-01

ABSTRACT Chromosomal copy number variation (CCNV) plays a key role in evolution and health of eukaryotes. The unicellular yeast Saccharomyces cerevisiae is an important model for studying the generation, physiological impact, and evolutionary significance of CCNV. Fundamental studies of this yeast have contributed to an extensive set of methods for analyzing and introducing CCNV. Moreover, these studies provided insight into the balance between negative and positive impacts of CCNV in evolutionary contexts. A growing body of evidence indicates that CCNV not only frequently occurs in industrial strains of Saccharomyces yeasts but also is a key contributor to the diversity of industrially relevant traits. This notion is further supported by the frequent involvement of CCNV in industrially relevant traits acquired during evolutionary engineering. This review describes recent developments in genome sequencing and genome editing techniques and discusses how these offer opportunities to unravel contributions of CCNV in industrial Saccharomyces strains as well as to rationally engineer yeast chromosomal copy numbers and karyotypes. PMID:28341679

Yeast Prions and Human Prion-like Proteins: Sequence Features and Prediction Methods

PubMed Central

Cascarina, Sean; Ross, Eric D.

2014-01-01

Prions are self-propagating infectious protein isoforms. A growing number of prions have been identified in yeast, each resulting from the conversion of soluble proteins into an insoluble amyloid form. These yeast prions have served as a powerful model system for studying the causes and consequences of prion aggregation. Remarkably, a number of human proteins containing prion-like domains, defined as domains with compositional similarity to yeast prion domains, have recently been linked to various human degenerative diseases, including amyotrophic lateral sclerosis (ALS). This suggests that the lessons learned from yeast prions may help in understanding these human diseases. In this review, we examine what has been learned about the amino acid sequence basis for prion aggregation in yeast, and how this information has been used to develop methods to predict aggregation propensity. We then discuss how this information is being applied to understand human disease, and the challenges involved in applying yeast prediction methods to higher organisms. PMID:24390581

Yeast prions and human prion-like proteins: sequence features and prediction methods.

PubMed

Cascarina, Sean M; Ross, Eric D

2014-06-01

Prions are self-propagating infectious protein isoforms. A growing number of prions have been identified in yeast, each resulting from the conversion of soluble proteins into an insoluble amyloid form. These yeast prions have served as a powerful model system for studying the causes and consequences of prion aggregation. Remarkably, a number of human proteins containing prion-like domains, defined as domains with compositional similarity to yeast prion domains, have recently been linked to various human degenerative diseases, including amyotrophic lateral sclerosis. This suggests that the lessons learned from yeast prions may help in understanding these human diseases. In this review, we examine what has been learned about the amino acid sequence basis for prion aggregation in yeast, and how this information has been used to develop methods to predict aggregation propensity. We then discuss how this information is being applied to understand human disease, and the challenges involved in applying yeast prediction methods to higher organisms.

The divergently transcribed genes encoding yeast ribosomal proteins L46 and S24 are activated by shared RPG-boxes.

PubMed Central

Kraakman, L S; Mager, W H; Maurer, K T; Nieuwint, R T; Planta, R J

1989-01-01

Transcription of the majority of the ribosomal protein (rp) genes in yeast is activated through common cis-acting elements, designated RPG-boxes. These elements have been shown to act as specific binding sites for the protein factor TUF/RAP1/GRF1 in vitro. Two such elements occur in the intergenic region separating the divergently transcribed genes encoding L46 and S24. To investigate whether the two RPG-boxes mediate transcription activation of both the L46 and S24 gene, two experimental strategies were followed: cloning of the respective genes on multicopy vectors and construction of fusion genes. Cloning of the L46 + S24 gene including the intergenic region in a multicopy yeast vector indicated that both genes are transcriptionally active. Using constructs in which only the S24 or the L46 gene is present, with or without the intergenic region, we obtained evidence that the intergenic region is indispensable for transcription activation of either gene. To demarcate the element(s) responsible for this activation, fusions of the intergenic region in either orientation to the galK reporter gene were made. Northern analysis of the levels of hybrid mRNA demonstrated that the intergenic region can serve as an heterologous promoter when it is in the 'S24-orientation'. Surprisingly, however, when fused in the reverse orientation the intergenic region did hardly confer transcription activity on the fusion gene. Furthermore, a 274 bp FnuDII-FnuDII fragment from the intergenic region that contains the RPG-boxes, could replace the naturally occurring upstream activation site (UASrpg) of the L25 rp-gene only when inserted in the 'S24-orientation'. Removal of 15 bp from the FnuDII fragment appeared to be sufficient to obtain transcription activation in the 'L46 orientation' as well. Analysis of a construct in which the RPG-boxes were selectively deleted from the promoter region of the L46 gene indicated that the RPG-boxes are needed for efficient transcriptional activation of the L46 gene. We conclude that all promoter elements for the S24 gene are located within the intergenic region, where the RPG-boxes are the most likely UAS-elements. However, the intergenic region (including the RPG-boxes) is required but not sufficient to confer transcription activity on the L46 gene. Images PMID:2602141

The divergently transcribed genes encoding yeast ribosomal proteins L46 and S24 are activated by shared RPG-boxes.

PubMed

Kraakman, L S; Mager, W H; Maurer, K T; Nieuwint, R T; Planta, R J

1989-12-11

Transcription of the majority of the ribosomal protein (rp) genes in yeast is activated through common cis-acting elements, designated RPG-boxes. These elements have been shown to act as specific binding sites for the protein factor TUF/RAP1/GRF1 in vitro. Two such elements occur in the intergenic region separating the divergently transcribed genes encoding L46 and S24. To investigate whether the two RPG-boxes mediate transcription activation of both the L46 and S24 gene, two experimental strategies were followed: cloning of the respective genes on multicopy vectors and construction of fusion genes. Cloning of the L46 + S24 gene including the intergenic region in a multicopy yeast vector indicated that both genes are transcriptionally active. Using constructs in which only the S24 or the L46 gene is present, with or without the intergenic region, we obtained evidence that the intergenic region is indispensable for transcription activation of either gene. To demarcate the element(s) responsible for this activation, fusions of the intergenic region in either orientation to the galK reporter gene were made. Northern analysis of the levels of hybrid mRNA demonstrated that the intergenic region can serve as an heterologous promoter when it is in the 'S24-orientation'. Surprisingly, however, when fused in the reverse orientation the intergenic region did hardly confer transcription activity on the fusion gene. Furthermore, a 274 bp FnuDII-FnuDII fragment from the intergenic region that contains the RPG-boxes, could replace the naturally occurring upstream activation site (UASrpg) of the L25 rp-gene only when inserted in the 'S24-orientation'. Removal of 15 bp from the FnuDII fragment appeared to be sufficient to obtain transcription activation in the 'L46 orientation' as well. Analysis of a construct in which the RPG-boxes were selectively deleted from the promoter region of the L46 gene indicated that the RPG-boxes are needed for efficient transcriptional activation of the L46 gene. We conclude that all promoter elements for the S24 gene are located within the intergenic region, where the RPG-boxes are the most likely UAS-elements. However, the intergenic region (including the RPG-boxes) is required but not sufficient to confer transcription activity on the L46 gene.

Reconstructing the Qo Site of Plasmodium falciparum bc 1 Complex in the Yeast Enzyme

PubMed Central

Vallières, Cindy; Fisher, Nicholas; Meunier, Brigitte

2013-01-01

The bc 1 complex of the mitochondrial respiratory chain is essential for Plasmodium falciparum proliferation, the causative agent of human malaria. Therefore, this enzyme is an attractive target for antimalarials. However, biochemical investigations of the parasite enzyme needed for the study of new drugs are challenging. In order to facilitate the study of new compounds targeting the enzyme, we are modifying the inhibitor binding sites of the yeast Saccharomyces cerevisiae to generate a complex that mimics the P. falciparum enzyme. In this study we focused on its Qo pocket, the site of atovaquone binding which is a leading antimalarial drug used in treatment and causal prophylaxis. We constructed and studied a series of mutants with modified Qo sites where yeast residues have been replaced by P. falciparum equivalents, or, for comparison, by human equivalents. Mitochondria were prepared from the yeast Plasmodium-like and human-like Qo mutants. We measured the bc 1 complex sensitivity to atovaquone, azoxystrobin, a Qo site targeting fungicide active against P. falciparum and RCQ06, a quinolone-derivative inhibitor of P. falciparum bc 1 complex.The data obtained highlighted variations in the Qo site that could explain the differences in inhibitor sensitivity between yeast, plasmodial and human enzymes. We showed that the yeast Plasmodium-like Qo mutants could be useful and easy-to-use tools for the study of that class of antimalarials. PMID:23951230

The SAGES Fundamental Use of Surgical Energy program (FUSE): history, development, and purpose.

PubMed

Fuchshuber, P; Schwaitzberg, S; Jones, D; Jones, S B; Feldman, L; Munro, M; Robinson, T; Purcell-Jackson, G; Mikami, D; Madani, A; Brunt, M; Dunkin, B; Gugliemi, C; Groah, L; Lim, R; Mischna, J; Voyles, C R

2018-06-01

Adverse events due to energy device use in surgical operating rooms are a daily occurrence. These occur at a rate of approximately 1-2 per 1000 operations. Hundreds of operating room fires occur each year in the United States, some causing severe injury and even mortality. The Society of American Gastrointestinal and Endoscopic Surgeons (SAGES) therefore created the first comprehensive educational curriculum on the safe use of surgical energy devices, called Fundamental Use of Surgical Energy (FUSE). This paper describes the history, development, and purpose of this important training program for all members of the operating room team. The databases of SAGES and the FUSE committee as well as personal photographs and documents of members of the FUSE task force were used to establish a brief history of the FUSE program from its inception to its current status. The authors were able to detail all aspects of the history, development, and national as well as global implementation of the third SAGES Fundamentals Program FUSE. The written documentation of the making of FUSE is an important contribution to the history and mission of SAGES and allows the reader to understand the idea, concept, realization, and implementation of the only free online educational tool for physicians on energy devices available today. FUSE is the culmination of the SAGES efforts to recognize gaps in patient safety and develop state-of-the-art educational programs to address those gaps. It is the goal of the FUSE task force to ensure that general FUSE implementation becomes multinational, involving as many countries as possible.

In Situ Analysis of Metabolic Characteristics Reveals the Key Yeast in the Spontaneous and Solid-State Fermentation Process of Chinese Light-Style Liquor

PubMed Central

Kong, Yu; Wu, Qun; Zhang, Yan

2014-01-01

The in situ metabolic characteristics of the yeasts involved in spontaneous fermentation process of Chinese light-style liquor are poorly understood. The covariation between metabolic profiles and yeast communities in Chinese light-style liquor was modeled using the partial least square (PLS) regression method. The diversity of yeast species was evaluated by sequence analysis of the 26S ribosomal DNA (rDNA) D1/D2 domains of cultivable yeasts, and the volatile compounds in fermented grains were analyzed by gas chromatography (GC)-mass spectrometry (MS). Eight yeast species and 58 volatile compounds were identified, respectively. The modulation of 16 of these volatile compounds was associated with variations in the yeast population (goodness of prediction [Q2] > 20%). The results showed that Pichia anomala was responsible for the characteristic aroma of Chinese liquor, through the regulation of several important volatile compounds, such as ethyl lactate, octanoic acid, and ethyl tetradecanoate. Correspondingly, almost all of the compounds associated with P. anomala were detected in a pure culture of this yeast. In contrast to the PLS regression results, however, ethyl lactate and ethyl isobutyrate were not detected in the same pure culture, which indicated that some metabolites could be generated by P. anomala only when it existed in a community with other yeast species. Furthermore, different yeast communities provided different volatile patterns in the fermented grains, which resulted in distinct flavor profiles in the resulting liquors. This study could help identify the key yeast species involved in spontaneous fermentation and provide a deeper understanding of the role of individual yeast species in the community. PMID:24727269

The effect of yeast (Saccharomyces cerevisiae) on nutrient intake, digestibility and finishing performance of lambs fed a diet based on dried molasses sugar beet-pulp.

PubMed

Payandeh, S; Kafilzadeh, F

2007-12-15

This experiment was conducted to determine the effect of yeast (Saccharomyces cerevisiae, SC47) on finishing performance, digestibility, some blood metabolites and carcass characteristics of male lambs fed a diet based on dried Molasses Sugar Beet-Pulp (MSBP). Eighteen Sanjabi male lambs (20.95 +/- 2.7 kg initial body weight and 3 month of age) were used in a completely randomized design. Animals were assigned to one of the two dietary treatments (with or without yeast). Digestibility and nitrogen balance experiment was carried out using six mature rams on finishing diet with and without yeast. Serum metabolites were determined in samples taken from lambs at the end of finishing period. Dry matter digestibility of finishing diet was significantly increased by yeast addition. However, yeast did not have any significant effect on apparent digestibility of OM, NDF, CP and energy. Nitrogen retention was also not affected by yeast addition. Yeast resulted in a significant increase in the average daily gain, dry matter and organic matter intake. However, feed conversion ratio was not significantly affected by addition of yeast. The concentration of the serum metabolites including glucose, urea, cholesterol, sodium, potassium, calcium, phosphorous and cratinine were not affected significantly by yeast supplementation, but triglyceride concentrations increased significantly when yeast was fed. Addition of yeast to the diet did not have any significant effect on the carcass characteristics. Results of this study suggest that feeding saccharomyces cerevisiae with a diet based on MSBP can improve the performance of fattening lambs without any change in carcass characteristics or cuts.

Changes in expression of oxidative stress related genes in grapefruit peel in response to yeast Metschnikowia fructicola

USDA-ARS?s Scientific Manuscript database

To gain insight into the mode of action of the yeast biocontrol agent, Metschnikowia fructicola, the transcription profiles of genes involved in oxidative stress were studied in grapefruit (Citrus paradis, 'Star Ruby') surface wounds following the application of the yeast antagonist. Three transcri...

Kazachstania gamospora and Wickerhamomyces subpelliculosus: Two alternative baker's yeasts in the modern bakery.

PubMed

Zhou, Nerve; Schifferdecker, Anna Judith; Gamero, Amparo; Compagno, Concetta; Boekhout, Teun; Piškur, Jure; Knecht, Wolfgang

2017-06-05

Saccharomyces cerevisiae, the conventional baker's yeast, remains the most domesticated yeast monopolizing the baking industry. Its rapid consumption of sugars and production of CO 2 are the most important attributes required to leaven the dough. New research attempts highlight that these attributes are not unique to S. cerevisiae, but also found in several non-conventional yeast species. A small number of these yeast species with similar properties have been described, but remain poorly studied. They present a vast untapped potential for the use as leavening agents and flavor producers due to their genetic and phylogenetic diversity. We assessed the potential of several non-conventional yeasts as leavening agents and flavor producers in dough-like conditions in the presence of high sugar concentrations and stressful environments mimicking conditions found in flour dough. We tested the capabilities of bread leavening and aroma formation in a microbread platform as well as in a bakery setup. Bread leavened with Kazachstania gamospora and Wickerhamomyces subpelliculosus had better overall results compared to control baker's yeast. In addition, both displayed higher stress tolerance and broader aroma profiles than the control baker's yeast. These attributes are important in bread and other farinaceous products, making K. gamospora and W. subpelliculosus highly applicable as alternative baker's yeasts. Copyright © 2017 Elsevier B.V. All rights reserved.

Past and Future of Non-Saccharomyces Yeasts: From Spoilage Microorganisms to Biotechnological Tools for Improving Wine Aroma Complexity

PubMed Central

Padilla, Beatriz; Gil, José V.; Manzanares, Paloma

2016-01-01

It is well established that non-Saccharomyces wine yeasts, considered in the past as undesired or spoilage yeasts, can enhance the analytical composition, and aroma profile of the wine. The contribution of non-Saccharomyces yeasts, including the ability to secret enzymes and produce secondary metabolites, glycerol and ethanol, release of mannoproteins or contributions to color stability, is species- and strain-specific, pointing out the key importance of a clever strain selection. The use of mixed starters of selected non-Saccharomyces yeasts with strains of Saccharomyces cerevisiae represents an alternative to both spontaneous and inoculated wine fermentations, taking advantage of the potential positive role that non-Saccharomyces wine yeast species play in the organoleptic characteristics of wine. In this context mixed starters can meet the growing demand for new and improved wine yeast strains adapted to different types and styles of wine. With the aim of presenting old and new evidences on the potential of non-Saccharomyces yeasts to address this market trend, we mainly review the studies focused on non-Saccharomyces strain selection and design of mixed starters directed to improve primary and secondary aroma of wines. The ability of non-Saccharomyces wine yeasts to produce enzymes and metabolites of oenological relevance is also discussed. PMID:27065975

Past and Future of Non-Saccharomyces Yeasts: From Spoilage Microorganisms to Biotechnological Tools for Improving Wine Aroma Complexity.

PubMed

Padilla, Beatriz; Gil, José V; Manzanares, Paloma

2016-01-01

It is well established that non-Saccharomyces wine yeasts, considered in the past as undesired or spoilage yeasts, can enhance the analytical composition, and aroma profile of the wine. The contribution of non-Saccharomyces yeasts, including the ability to secret enzymes and produce secondary metabolites, glycerol and ethanol, release of mannoproteins or contributions to color stability, is species- and strain-specific, pointing out the key importance of a clever strain selection. The use of mixed starters of selected non-Saccharomyces yeasts with strains of Saccharomyces cerevisiae represents an alternative to both spontaneous and inoculated wine fermentations, taking advantage of the potential positive role that non-Saccharomyces wine yeast species play in the organoleptic characteristics of wine. In this context mixed starters can meet the growing demand for new and improved wine yeast strains adapted to different types and styles of wine. With the aim of presenting old and new evidences on the potential of non-Saccharomyces yeasts to address this market trend, we mainly review the studies focused on non-Saccharomyces strain selection and design of mixed starters directed to improve primary and secondary aroma of wines. The ability of non-Saccharomyces wine yeasts to produce enzymes and metabolites of oenological relevance is also discussed.

Specialist nectar-yeasts decline with urbanization in Berlin

NASA Astrophysics Data System (ADS)

Wehner, Jeannine; Mittelbach, Moritz; Rillig, Matthias C.; Verbruggen, Erik

2017-03-01

Nectar yeasts are common inhabitants of insect-pollinated flowers but factors determining their distribution are not well understood. We studied the influence of host identity, environmental factors related to pollution/urbanization, and the distance to a target beehive on local distribution of nectar yeasts within Robinia pseudoacacia L. and Tilia tomentosa Moench in Berlin, Germany. Nectar samples of six individuals per species were collected at seven sites in a 2 km radius from each target beehive and plated on YM-Agar to visualise the different morphotypes, which were then identified by sequencing a section of the 26S rDNA gene. Multivariate linear models were used to analyze the effects of all investigated factors on yeast occurrence per tree. Yeast distribution was mainly driven by host identity. The influence of the environmental factors (NO2, height of construction, soil sealing) strongly depended on the radius around the tree, similar to the distance of the sampled beehive. Incidence of specialist nectar-borne yeast species decreased with increasing pollution/urbanization index. Given that specialist yeast species gave way to generalist yeasts that have a reduced dependency on pollinators for between-flower dispersal, our results indicate that increased urbanization may restrict the movement of nectar-specialized yeasts, via limitations of pollinator foraging behavior.

A Simple and Rapid Protocol for Producing Yeast Extract from Saccharomyces cerevisiae Suitable for Preparing Bacterial Culture Media

PubMed Central

Zarei, Omid; Dastmalchi, Siavoush; Hamzeh-Mivehroud, Maryam

2016-01-01

Yeasts, especially Saccharomyces cerevisiae, are one of the oldest organisms with broad spectrum of applications, owing to their unique genetics and physiology. Yeast extract, i.e. the product of yeast cells, is extensively used as nutritional resource in bacterial culture media. The aim of this study was to develop a simple, rapid and cost benefit process to produce the yeast extract. In this procedure mechanical methods such as high temperature and pressure were utilized to produce the yeast extract. The growth of the bacteria feed with the produced yeast extract was monitored in order to assess the quality of the product. The results showed that the quality of the produced yeast extract was very promising concluded from the growth pattern of bacterial cells in media prepared from this product and was comparable with that of the three commercial yeast extracts in terms of bacterial growth properties. One of the main advantages of the current method was that no chemicals and enzymes were used, leading to the reduced production cost. The method is very simple and cost effective, and can be performed in a reasonable time making it suitable for being adopted by research laboratories. Furthermore, it can be scaled up to produce large quantities for industrial applications. PMID:28243289

Enhanced leavening properties of baker's yeast by reducing sucrase activity in sweet dough.

PubMed

Zhang, Cui-Ying; Lin, Xue; Feng, Bing; Liu, Xiao-Er; Bai, Xiao-Wen; Xu, Jia; Pi, Li; Xiao, Dong-Guang

2016-07-01

Leavening ability in sweet dough is required for the commercial applications of baker's yeast. This property depends on many factors, such as glycolytic activity, sucrase activity, and osmotolerance. This study explored the importance of sucrase level on the leavening ability of baker's yeast in sweet dough. Furthermore, the baker's yeast strains with varying sucrase activities were constructed by deleting SUC2, which encodes sucrase or replacing the SUC2 promoter with the VPS8/TEF1 promoter. The results verify that the sucrase activity negatively affects the leavening ability of baker's yeast strains under high-sucrose conditions. Based on a certain level of osmotolerance, sucrase level plays a significant role in the fermentation performance of baker's yeast, and appropriate sucrase activity is an important determinant for the leavening property of baker's yeast in sweet dough. Therefore, modification on sucrase activity is an effective method for improving the leavening properties of baker's yeast in sweet dough. This finding provides guidance for the breeding of industrial baker's yeast strains for sweet dough leavening. The transformants BS1 with deleted SUC2 genetic background provided decreased sucrase activity (a decrease of 39.3 %) and exhibited enhanced leavening property (an increase of 12.4 %). Such a strain could be useful for industrial applications.

Enumeration and rapid identification of yeasts during extraction processes of extra virgin olive oil in Tuscany.

PubMed

Mari, Eleonora; Guerrini, Simona; Granchi, Lisa; Vincenzini, Massimo

2016-06-01

The aim of this study was to evaluate the occurrence of yeast populations during different olive oil extraction processes, carried out in three consecutive years in Tuscany (Italy), by analysing crushed pastes, kneaded pastes, oil from decanter and pomaces. The results showed yeast concentrations ranging between 10(3) and 10(5) CFU/g or per mL. Seventeen dominant yeast species were identified by random amplified polymorphic DNA with primer M13 and their identification was confirmed by restriction fragments length polymorphism of ribosomal internal transcribed spacer and sequencing rRNA genes. The isolation frequencies of each species in the collected samples pointed out that the occurrence of the various yeast species in olive oil extraction process was dependent not only on the yeasts contaminating the olives but also on the yeasts colonizing the plant for oil extraction. In fact, eleven dominant yeast species were detected from the washed olives, but only three of them were also found in oil samples at significant isolation frequency. On the contrary, the most abundant species in oil samples, Yamadazyma terventina, did not occur in washed olive samples. These findings suggest a phenomenon of contamination of the plant for oil extraction that selects some yeast species that could affect the quality of olive oil.

Virgin olive oil yeasts: A review.

PubMed

Ciafardini, Gino; Zullo, Biagi Angelo

2018-04-01

This review summarizes current knowledge on virgin olive oil yeasts. Newly produced olive oil contains solid particles and micro drops of vegetation water in which yeasts reproduce to become the typical microbiota of olive oil. To date, about seventeen yeast species have been isolated from different types of olive oils and their by-products, of which six species have been identified as new species. Certain yeast species contribute greatly to improving the sensorial characteristics of the newly produced olive oil, whereas other species are considered harmful as they can damage the oil quality through the production of unpleasant flavors and triacylglycerol hydrolysis. Studies carried out in certain yeast strains have demonstrated the presence of defects in olive oil treated with Candida adriatica, Nakazawaea wickerhamii and Candida diddensiae specific strains, while other olive oil samples treated with other Candida diddensiae strains were defect-free after four months of storage and categorized as extra virgin. A new acetic acid producing yeast species, namely, Brettanomyces acidodurans sp. nov., which was recently isolated from olive oil, could be implicated in the wine-vinegary defect of the product. Other aspects related to the activity of the lipase-producing yeasts and the survival of the yeast species in the flavored olive oils are also discussed. Copyright © 2017 Elsevier Ltd. All rights reserved.

Concentration-Dependent Effects of Rhodiola Rosea on Long-Term Survival and Stress Resistance of Yeast Saccharomyces Cerevisiae: The Involvement of YAP 1 and MSN2/4 Regulatory Proteins

PubMed Central

Bayliak, Maria M.; Burdyliuk, Nadia I.; Izers’ka, Lilia I.; Lushchak, Volodymyr I.

2014-01-01

Concentration-dependent effects of aqueous extract from R. rosea root on long-term survival and stress resistance of budding yeast Saccharomyces cerevisiae were studied. At low concentrations, R. rosea aqueous extract extended yeast chronological lifespan, enhanced oxidative stress resistance of stationary-phase cells and resistance to number stressors in exponentially growing cultures. At high concentrations, R. rosea extract sensitized yeast cells to stresses and shortened yeast lifespan. These biphasic concentration-responses describe a common hormetic phenomenon characterized by a low-dose stimulation and a high-dose inhibition. Yeast pretreatment with low doses of R. rosea extract enhanced yeast survival and prevented protein oxidation under H2O2-induced oxidative stress. Positive effect of R. rosea extract on yeast survival under heat shock exposure was not accompanied with changes in antioxidant enzyme activities and levels of oxidized proteins. The deficiency in transcriptional regulators, Msn2/Msn4 and Yap1, abolished the positive effect of low doses of R. rosea extract on yeast viability under stress challenges. Potential involvement of Msn2/Msn4 and Yap1 regulatory proteins in realization of R. rosea beneficial effects is discussed. PMID:24659935

Yeasts and yeast-like organisms associated with fruits and blossoms of different fruit trees.

PubMed

Vadkertiová, Renáta; Molnárová, Jana; Vránová, Dana; Sláviková, Elena

2012-12-01

Yeasts are common inhabitants of the phyllosphere, but our knowledge of their diversity in various plant organs is still limited. This study focused on the diversity of yeasts and yeast-like organisms associated with matured fruits and fully open blossoms of apple, plum, and pear trees, during 2 consecutive years at 3 localities in southwest Slovakia. The occurrence of yeasts and yeast-like organisms in fruit samples was 2½ times higher and the yeast community more diverse than that in blossom samples. Only 2 species (Aureobasidium pullulans and Metschnikowia pulcherrima) occurred regularly in the blossom samples, whereas Galactomyces candidus, Hanseniaspora guilliermondii, Hanseniaspora uvarum, M. pulcherrima, Pichia kluyveri, Pichia kudriavzevii, and Saccharomyces cerevisiae were the most frequently isolated species from the fruit samples. The ratio of the number of samples where only individual species were present to the number of samples where 2 or more species were found (consortium) was counted. The occurrence of individual species in comparison with consortia was much higher in blossom samples than in fruit samples. In the latter, consortia predominated. Aureobasidium pullulans, M. pulcherrima, and S. cerevisiae, isolated from both the fruits and blossoms, can be considered as resident yeast species of various fruit tree species cultivated in southwest Slovakia localities.

Effect of Yeast Cell Morphology, Cell Wall Physical Structure and Chemical Composition on Patulin Adsorption

PubMed Central

Luo, Ying; Wang, Jianguo; Liu, Bin; Wang, Zhouli; Yuan, Yahong; Yue, Tianli

2015-01-01

The capability of yeast to adsorb patulin in fruit juice can aid in substantially reducing the patulin toxic effect on human health. This study aimed to investigate the capability of yeast cell morphology and cell wall internal structure and composition to adsorb patulin. To compare different yeast cell morphologies, cell wall internal structure and composition, scanning electron microscope, transmission electron microscope and ion chromatography were used. The results indicated that patulin adsorption capability of yeast was influenced by cell surface areas, volume, and cell wall thickness, as well as 1,3-β-glucan content. Among these factors, cell wall thickness and 1,3-β-glucan content serve significant functions. The investigation revealed that patulin adsorption capability was mainly affected by the three-dimensional network structure of the cell wall composed of 1,3-β-glucan. Finally, patulin adsorption in commercial kiwi fruit juice was investigated, and the results indicated that yeast cells could adsorb patulin from commercial kiwi fruit juice efficiently. This study can potentially simulate in vitro cell walls to enhance patulin adsorption capability and successfully apply to fruit juice industry. PMID:26295574

Effect of Yeast Cell Morphology, Cell Wall Physical Structure and Chemical Composition on Patulin Adsorption.

PubMed

Luo, Ying; Wang, Jianguo; Liu, Bin; Wang, Zhouli; Yuan, Yahong; Yue, Tianli

2015-01-01

The capability of yeast to adsorb patulin in fruit juice can aid in substantially reducing the patulin toxic effect on human health. This study aimed to investigate the capability of yeast cell morphology and cell wall internal structure and composition to adsorb patulin. To compare different yeast cell morphologies, cell wall internal structure and composition, scanning electron microscope, transmission electron microscope and ion chromatography were used. The results indicated that patulin adsorption capability of yeast was influenced by cell surface areas, volume, and cell wall thickness, as well as 1,3-β-glucan content. Among these factors, cell wall thickness and 1,3-β-glucan content serve significant functions. The investigation revealed that patulin adsorption capability was mainly affected by the three-dimensional network structure of the cell wall composed of 1,3-β-glucan. Finally, patulin adsorption in commercial kiwi fruit juice was investigated, and the results indicated that yeast cells could adsorb patulin from commercial kiwi fruit juice efficiently. This study can potentially simulate in vitro cell walls to enhance patulin adsorption capability and successfully apply to fruit juice industry.

Sensitivity of fructose-1,6-biphosphatase from yeast, liver and skeletal muscle to fructose-2,6-biphosphate and 5'-adenosine monophosphate.

PubMed

von Herrath, M; Holzer, H

1988-05-01

As a prerequisite for future studies on the possible effect of sulphite, an anti-microbial agent, on gluconeogenesis in yeast, a comparative study of fructose-1,6-bisphosphatase (FBPase), a key enzyme of gluconeogenesis, from yeast, liver and skeletal muscle is reported. In contrast to FBPase from yeast or liver, FBPase from skeletal muscle is approximately 1000-fold more sensitive to inhibition by 5' adenosine monophosphate and 30 to 250-fold less sensitive to inhibition by fructose-2,6-bisphosphate. The kinetic properties of the FBPases, determined by the ratios R(Mg2+/Mn2+) and R (pH 7/9) of the enzyme activities, measured at 10 mM Mg2+ and 2 mM Mn2+ and at pH 7.0 and 9.0, respectively, show a drastic difference between the skeletal muscle and the yeast or liver enzymes. The data support the idea that the enzymes from yeast and liver function in gluconeogenesis, whereas the enzyme from skeletal muscle is involved in other biological functions.

The Yeast Copper Response Is Regulated by DNA Damage

PubMed Central

Dong, Kangzhen; Addinall, Stephen G.; Lydall, David

2013-01-01

Copper is an essential but potentially toxic redox-active metal, so the levels and distribution of this metal are carefully regulated to ensure that it binds to the correct proteins. Previous studies of copper-dependent transcription in the yeast Saccharomyces cerevisiae have focused on the response of genes to changes in the exogenous levels of copper. We now report that yeast copper genes are regulated in response to the DNA-damaging agents methyl methanesulfonate (MMS) and hydroxyurea by a mechanism(s) that requires the copper-responsive transcription factors Mac1 and AceI, copper superoxide dismutase (Sod1) activity, and the Rad53 checkpoint kinase. Furthermore, in copper-starved yeast, the response of the Rad53 pathway to MMS is compromised due to a loss of Sod1 activity, consistent with the model that yeast imports copper to ensure Sod1 activity and Rad53 signaling. Crucially, the Mac1 transcription factor undergoes changes in its redox state in response to changing levels of copper or MMS. This study has therefore identified a novel regulatory relationship between cellular redox, copper homeostasis, and the DNA damage response in yeast. PMID:23959798

Diversity of soil yeasts isolated from South Victoria Land, Antarctica

USGS Publications Warehouse

Connell, L.; Redman, R.; Craig, S.; Scorzetti, G.; Iszard, M.; Rodriguez, R.

2008-01-01

Unicellular fungi, commonly referred to as yeasts, were found to be components of the culturable soil fungal population in Taylor Valley, Mt. Discovery, Wright Valley, and two mountain peaks of South Victoria Land, Antarctica. Samples were taken from sites spanning a diversity of soil habitats that were not directly associated with vertebrate activity. A large proportion of yeasts isolated in this study were basidiomycetous species (89%), of which 43% may represent undescribed species, demonstrating that culturable yeasts remain incompletely described in these polar desert soils. Cryptococcus species represented the most often isolated genus (33%) followed by Leucosporidium (22%). Principle component analysis and multiple linear regression using stepwise selection was used to model the relation between abiotic variables (principle component 1 and principle component 2 scores) and yeast biodiversity (the number of species present at a given site). These analyses identified soil pH and electrical conductivity as significant predictors of yeast biodiversity. Species-specific PCR primers were designed to rapidly discriminate among the Dioszegia and Leucosporidium species collected in this study. ?? 2008 Springer Science+Business Media, LLC.

SEM study of the effects of bacteria and yeasts on wood decay by brown and white-rot fungi. [Enterobacter, Cryptococcus Pichia, and Saccharomyces

DOE Office of Scientific and Technical Information (OSTI.GOV)

Blanchette, R.A.; Shaw, C.G.; Cohen, A.L.

The scanning electron microscope was used to 1) examine the associations among microorganisms during wood decay and 2) observe the effect of these organisms on degradation of cell wall components. Bacteria (Enterobacter) and yeasts (Cryptococcus Pichia, and Saccharomyces) were found to have a mutualistic association with a white-rot fungus during decay of coniferous wood. Coriolus (Polyporus versicolar) degraded cell wall components in a typical ''erosion trough'' manner (i.e., by lysing zones around fungal hyphae). Bacteria and yeasts were seen only in these lysed zones. Typical gross decay patterns caused by the white-rot fungus were unaltered by bacteria and yeasts. Themore » SEM study suggests that the decay process is enhanced when these organisms are associated. In contrast, the same bacteria and yeasts were inhibitory when combined with a brown-rot fungus.« less

Exploring the northern limit of the distribution of Saccharomyces cerevisiae and Saccharomyces paradoxus in North America.

PubMed

Charron, Guillaume; Leducq, Jean-Baptiste; Bertin, Chloé; Dubé, Alexandre K; Landry, Christian R

2014-03-01

We examined the northern limit of Saccharomyces cerevisiae and Saccharomyces paradoxus in northeast America. We collected 876 natural samples at 29 sites and applied enrichment methods for the isolation of mesophilic yeasts. We uncovered a large diversity of yeasts, in some cases, associated with specific substrates. Sequencing of the ITS1, 5.8S and ITS2 loci allowed to assign 226 yeast strains at the species level, including 41 S. paradoxus strains. Our intensive sampling suggests that if present, S. cerevisiae is rare at these northern latitudes. Our sampling efforts spread across several months of the year revealed that successful sampling increases throughout the summer and diminishes significantly at the beginning of the fall. The data obtained on the ecological context of yeasts corroborate what was previously reported on Pichiaceae, Saccharomycodaceae, Debaryomycetaceae and Phaffomycetaceae yeast families. We identified 24 yeast isolates that could not be assigned to any known species and that may be of taxonomic, medical, or biotechnological importance. Our study reports new data on the taxonomic diversity of yeasts and new resources for studying the evolution and ecology of S. paradoxus. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Genetic analysis of Saccharomyces cerevisiae strains isolated from palm wine in eastern Nigeria. Comparison with other African strains.

PubMed

Ezeronye, O U; Legras, J-L

2009-05-01

To study the yeast diversity of Nigerian palm wines by comparison with other African strains. Twenty-three Saccharomyces cerevisiae strains were obtained from palm wine samples collected at four locations in eastern Nigeria, and characterized using different molecular techniques: internal transcribed spacer restriction fragment length polymorphism and sequence analysis, pulsed field gel electrophoresis, inter delta typing and microsatellite multilocus analysis. These techniques revealed that palm wine yeasts represent a group of closely related strains that includes other West African isolates (CBS400, NCYC110, DVPG6044). Population analysis revealed an excess of homozygote strains and an allelic richness similar to wine suggestive of local domestication. Several other African yeast strains were not connected to this group. Ghana sorghum beer strains and other African strains (DBVPG1853 and MUCL28071) displayed strikingly high relatedness with European bread, beer or wine strains, and the genome of strain MUCL30909 contained African and wine-type alleles, indicating its hybrid origin. Nigerian palm wine yeast represents a local specific yeast flora, whereas a European origin or hybrid was suspected for several other Africa isolates. This study presents the first genetic characterization of an autochthonous African palm wine yeast population and confirms the idea that human intervention has favoured yeast migration.

[Effects of laser welding on bond of porcelain fused cast pure titanium].

PubMed

Zhu, Juan-fang; He, Hui-ming; Gao, Bo; Wang, Zhong-yi

2006-04-01

To investigate the influence of the laser welding on bond of porcelain fused to cast pure titanium. Twenty cast titanium plates were divided into two groups: laser welded group and control group. The low-fusing porcelain was fused to the laser welded cast pure titanium plates at fusion zone. The bond strength of the porcelain to laser welded cast pure titanium was measured by the three-point bending test. The interface of titanium and porcelain was investigated by scanning electron microscopy (SEM) and energy depressive X-ray detector (EDX). The non-welded titanium plates were used as comparison. No significant difference of the bond strength was found between laser-welded samples [(46.85 +/- 0.76) MPa] and the controls [(41.71 +/- 0.55) MPa] (P > 0.05). The SEM displayed the interface presented similar irregularities with a predominance. The titanium diffused to low-fusing porcelain, while silicon and aluminum diffused to titanium basement. Laser welding does not affect low-fusing porcelain fused to pure titanium.

Saccharomyces cerevisiae as a model for the study of extranuclear functions of mammalian telomerase.

PubMed

Simonicova, Lucia; Dudekova, Henrieta; Ferenc, Jaroslav; Prochazkova, Katarina; Nebohacova, Martina; Dusinsky, Roman; Nosek, Jozef; Tomaska, Lubomir

2015-11-01

The experimental evidence from the last decade made telomerase a prominent member of a family of moonlighting proteins performing different functions at various cellular loci. However, the study of extratelomeric functions of the catalytic subunit of mammalian telomerase (TERT) is often complicated by the fact that it is sometimes difficult to distinguish them from its role(s) at the chromosomal ends. Here, we present an experimental model for studying the extranuclear function(s) of mammalian telomerase in the yeast Saccharomyces cerevisiae. We demonstrate that the catalytic subunit of mammalian telomerase protects the yeast cells against oxidative stress and affects the stability of the mitochondrial genome. The advantage of using S. cerevisiae to study of mammalian telomerase is that (1) mammalian TERT does not interfere with its yeast counterpart in the maintenance of telomeres, (2) yeast telomerase is not localized in mitochondria and (3) it does not seem to be involved in the protection of cells against oxidative stress and stabilization of mtDNA. Thus, yeast cells can be used as a 'test tube' for reconstitution of mammalian TERT extranuclear function(s).

Improvement of tolerance to freeze-thaw stress of baker's yeast by cultivation with soy peptides.

PubMed

Izawa, Shingo; Ikeda, Kayo; Takahashi, Nobuyuki; Inoue, Yoshiharu

2007-06-01

The tolerance to freeze-thaw stress of yeast cells is critical for frozen-dough technology in the baking industry. In this study, we examined the effects of soy peptides on the freeze-thaw stress tolerance of yeast cells. We found that the cells cultured with soy peptides acquired improved tolerance to freeze-thaw stress and retained high leavening ability in dough after frozen storage for 7 days. The final quality of bread regarding its volume and texture was also improved by using yeast cells cultured with soy peptides. These findings promote the utilization of soy peptides as ingredients of culture media to improve the quality of baker's yeast.

Yeast and Macroinvertebrate Communities Associated with Leaf Litter Decomposition in a Second Order Stream

NASA Astrophysics Data System (ADS)

Sampaio, Ana; Cortes, Rui; Leão, Cecília

2004-11-01

The composition of yeast and macroinvertebrate communities was studied on black alder, blue gum eucalyptus and English oak leaves decaying in a stream during a six-month period. ANOVA analysis showed significantly different values (p < 0.0001) of yeast and macroinvertebrate densities among the three leaf litters. Some yeast species such as Cryptococcus albidus (Saito), C. laurentii (Kufferath), Rhodothorula glutinis (Fresenius), R. colostri (Castelli), and Debaryomyces hansenii (Lodder and Kreger-van Rij) were present in all litter types. Other yeasts were restricted to a specific type of litter. Macroinvertebrates were dominated by collectors-gatherers on oak and eucalyptus leaves. Shredders reached highest densities in alder leaves. (

Oxidative Stress and Programmed Cell Death in Yeast

PubMed Central

Farrugia, Gianluca; Balzan, Rena

2012-01-01

Yeasts, such as Saccharomyces cerevisiae, have long served as useful models for the study of oxidative stress, an event associated with cell death and severe human pathologies. This review will discuss oxidative stress in yeast, in terms of sources of reactive oxygen species (ROS), their molecular targets, and the metabolic responses elicited by cellular ROS accumulation. Responses of yeast to accumulated ROS include upregulation of antioxidants mediated by complex transcriptional changes, activation of pro-survival pathways such as mitophagy, and programmed cell death (PCD) which, apart from apoptosis, includes pathways such as autophagy and necrosis, a form of cell death long considered accidental and uncoordinated. The role of ROS in yeast aging will also be discussed. PMID:22737670

Genome Diversity and Evolution in the Budding Yeasts (Saccharomycotina)

PubMed Central

Dujon, Bernard A.; Louis, Edward J.

2017-01-01

Considerable progress in our understanding of yeast genomes and their evolution has been made over the last decade with the sequencing, analysis, and comparisons of numerous species, strains, or isolates of diverse origins. The role played by yeasts in natural environments as well as in artificial manufactures, combined with the importance of some species as model experimental systems sustained this effort. At the same time, their enormous evolutionary diversity (there are yeast species in every subphylum of Dikarya) sparked curiosity but necessitated further efforts to obtain appropriate reference genomes. Today, yeast genomes have been very informative about basic mechanisms of evolution, speciation, hybridization, domestication, as well as about the molecular machineries underlying them. They are also irreplaceable to investigate in detail the complex relationship between genotypes and phenotypes with both theoretical and practical implications. This review examines these questions at two distinct levels offered by the broad evolutionary range of yeasts: inside the best-studied Saccharomyces species complex, and across the entire and diversified subphylum of Saccharomycotina. While obviously revealing evolutionary histories at different scales, data converge to a remarkably coherent picture in which one can estimate the relative importance of intrinsic genome dynamics, including gene birth and loss, vs. horizontal genetic accidents in the making of populations. The facility with which novel yeast genomes can now be studied, combined with the already numerous available reference genomes, offer privileged perspectives to further examine these fundamental biological questions using yeasts both as eukaryotic models and as fungi of practical importance. PMID:28592505

Yeasts as important agents of onychomycosis: in vitro activity of propolis against yeasts isolated from patients with nail infection.

PubMed

Khosravi, Ali Reza; Shokri, Hojjatollah; Nikaein, Donya; Mansouri, Parvin; Erfanmanesh, Ahmad; Chalangari, Reza; Katalin, Martis

2013-01-01

The purposes of this study were to determine the frequency of the yeast species obtained from patients with clinical features of onychomycosis and the in vitro antifungal susceptibility of the yeast species to propolis. A prospective study was carried out at the Mycology Research Center in Iran from 2010 to 2011. Clinical diagnosis was performed by direct microscopic examination and culture. Different yeast species were identified by morphological and biochemical tests. An antifungal susceptibility test to fluconazole (FLU) and propolis by the broth microdilution method was performed on each isolate. One hundred and twenty-eight fungal isolates were obtained. The most prevalent fungi were yeasts (81, 63.2%), dermatophytes (36, 28.1%), and nondermatophyte fungi (11, 8.6%). Fingernails were more affected than toenails (65.4% vs. 19.8%, respectively). The most frequently found species was Candida albicans (38.5%), followed by Candida spp. (23.1%), C. tropicalis (10.8%), C. kefyr (6.2%), C. krusei (3.1%), Malassezia globosa (4.6%), M. slooffiae (4.6%), and M. pachydermatis (1.5%). Of all yeast isolates (65), seven showed resistance to FLU. The average MIC of propolis for FLU-susceptible isolates was 5.8 μg/mL, whereas this value was 12.25 μg/mL for FLU-resistant isolates. Our results proved that the propolis inhibits the growth of pathogenic yeasts and confirmed the efficiency of propolis as an anti-Candida and anti-Malassezia agent.

Microbial-enhanced lindane removal by sugarcane (Saccharum officinarum) in doped soil-applications in phytoremediation and bioaugmentation.

PubMed

Salam, Jaseetha Abdul; Hatha, Mohammed A A; Das, Nilanjana

2017-05-15

The aim of this study was to examine the effect of lindane-degrading yeast on the growth and lindane uptake by Saccharum sp., in doped garden soils. The rhizosphere of Saccharum plant was amended with yeast Candida VITJzN04 by root-inoculation. The bio-augment yeast was applied in two different forms viz., planktonic form and cells immobilized on sugarcane-bagasse, in the pot experiments. Garden soils (lindane∼100 mg/kg) exposed to various treatments were monitored for a period of 30 days, for residual lindane by gas-chromatography analysis. The lindane-removal rates in soil were expressed in terms of half-life period and were recorded as 13.3 days (yeast), 43.3 days (Saccharum), 9.8 days (free yeast-plant) and 7.1 days (immobilized yeast-plant). Additionally, Candida sp., was also identified as a plant growth promoting yeast due to its ability to produce growth hormone and solubilize insoluble phosphates in the soil for better uptake by the plant species. Bio-stimulation of the soil with yeast immobilized on sugarcane bagasse further enhanced the total yeast activity in the soil which in turn had a positive influence on lindane-removal. Combined treatment with bagasse immobilized yeast and plant showed the best lindane degradation. Results suggested that the synergistic activity of plant and yeast resulted in fast and efficient degradation of lindane. Thus, it can be concluded that Saccharum plant in combination with Candida VITJzN04 is an effective alternative for the conventional remediation strategies. Copyright © 2017 Elsevier Ltd. All rights reserved.

Electron transport chain in a thermotolerant yeast.

PubMed

Mejía-Barajas, Jorge A; Martínez-Mora, José A; Salgado-Garciglia, Rafael; Noriega-Cisneros, Ruth; Ortiz-Avila, Omar; Cortés-Rojo, Christian; Saavedra-Molina, Alfredo

2017-04-01

Yeasts capable of growing and surviving at high temperatures are regarded as thermotolerant. For appropriate functioning of cellular processes and cell survival, the maintenance of an optimal redox state is critical of reducing and oxidizing species. We studied mitochondrial functions of the thermotolerant Kluyveromyces marxianus SLP1 and the mesophilic OFF1 yeasts, through the evaluation of its mitochondrial membrane potential (ΔΨ m ), ATPase activity, electron transport chain (ETC) activities, alternative oxidase activity, lipid peroxidation. Mitochondrial membrane potential and the cytoplasmic free Ca 2+ ions (Ca 2+ cyt) increased in the SLP1 yeast when exposed to high temperature, compared with the mesophilic yeast OFF1. ATPase activity in the mesophilic yeast diminished 80% when exposed to 40° while the thermotolerant SLP1 showed no change, despite an increase in the mitochondrial lipid peroxidation. The SLP1 thermotolerant yeast exposed to high temperature showed a diminution of 33% of the oxygen consumption in state 4. The uncoupled state 3 of oxygen consumption did not change in the mesophilic yeast when it had an increase of temperature, whereas in the thermotolerant SLP1 yeast resulted in an increase of 2.5 times when yeast were grown at 30 o , while a decrease of 51% was observed when it was exposed to high temperature. The activities of the ETC complexes were diminished in the SLP1 when exposed to high temperature, but also it was distinguished an alternative oxidase activity. Our results suggest that the mitochondria state, particularly ETC state, is an important characteristic of the thermotolerance of the SLP1 yeast strain.

Extracellular enzymatic activities and physiological profiles of yeasts colonizing fruit trees.

PubMed

Molnárová, Jana; Vadkertiová, Renáta; Stratilová, Eva

2014-07-01

Yeasts form a significant and diverse part of the phyllosphere microbiota. Some yeasts that inhabit plants have been found to exhibit extracellular enzymatic activities. The aim of the present study was to investigate the ability of yeasts isolated from leaves, fruits, and blossoms of fruit trees cultivated in Southwest Slovakia to produce extracellular enzymes, and to discover whether the yeasts originating from these plant organs differ from each other in their physiological properties. In total, 92 strains belonging to 29 different species were tested for: extracellular protease, β-glucosidase, lipase, and polygalacturonase activities; fermentation abilities; the assimilation of xylose, saccharose and alcohols (methanol, ethanol, glycerol); and for growth in a medium with 33% glucose. The black yeast Aureobasidium pullulans showed the largest spectrum of activities of all the species tested. Almost 70% of the strains tested demonstrated some enzymatic activity, and more than 90% utilized one of the carbon compounds tested. Intraspecies variations were found for the species of the genera Cryptococcus and Pseudozyma. Interspecies differences of strains exhibiting some enzymatic activities and utilizing alcohols were also noted. The largest proportion of the yeasts exhibited β-glucosidase activity and assimilated alcohols independently of their origin. The highest number of strains positive for all activities tested was found among the yeasts associated with leaves. Yeasts isolated from blossoms assimilated saccharose and D-xylose the most frequently of all the yeasts tested. The majority of the fruit-inhabiting yeasts grew in the medium with higher osmotic pressure. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Effect of yeast supplementation on performance, rumination time, and rumen pH of dairy cows in commercial farm environments.

PubMed

Ambriz-Vilchis, V; Jessop, N S; Fawcett, R H; Webster, M; Shaw, D J; Walker, N; Macrae, A I

2017-07-01

The aims of the present study were to evaluate the effects of live yeast supplementation (Vistacell MUCL 39855, AB Vista, Marlborough, UK) on performance, rumination time, and rumen pH on dairy cows in commercial farm environments. Three trials were carried out, the trials lasted 12 (trial 1), 15 (trial 2), and 19 wk (trial 3). In each trial, 14 multiparous Holstein dairy cows were allocated to 2 groups that received (trial 1) a standard diet plus yeast, (trial 2) an acidogenic diet plus yeast, and (trial 3) grazing pasture plus yeast. Milk production, milk chemical characteristics, body weight and body condition score, rumination time, and rumen pH were monitored for each group throughout the 3 trials. No statistically significant differences were observed in any of the 3 trials for any of the recorded variables. In contrast, an effect of time (period or days in milk) on rumen pH was observed in all 3 of the trials, as time spent under the acidotic thresholds increased across the experimental periods; however the differences were not associated with live yeast supplementation. No effect of live yeast supplementation was observed in any of the 3 trials reported. Further research should include studies on animals at different stages of lactation (with emphasis on transition period and early lactation), consuming more challenging diets (higher level of inclusion of concentrates or starch), or under different environments such as grazing of succulent forages. Such studies might be required to elucidate any possible effect of live yeast supplementation of dairy cows when the rumen environment is under challenge. Copyright © 2017 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Internal fuse modules for solid tantalum capacitors

NASA Technical Reports Server (NTRS)

Dematos, H. V.

1981-01-01

Miniature fuse modules were designed for and incorporated into two styles of solid tantalum capacitors. One is an epoxy molded, radial leaded, high frequency decoupling capacitor; the other is an hermetically sealed device with axial lead wires. The fusible element for both devices consists of a fine bimetallic wire which reacts exothermically upon reaching a critical temperature and then disintegrates. The desirability of having fused devices is discussed and design constraints, in particular those which minimize inductance and series resistance while optimizing fuse actuation characteristics, are reviewed. Factors affecting the amount of energy required to actuate the fuse and reliability of acuation are identified.

Beginnings of microbiology and biochemistry: the contribution of yeast research.

PubMed

Barnett, James A

2003-03-01

With improvements in microscopes early in the nineteenth century, yeasts were seen to be living organisms, although some famous scientists ridiculed the idea and their influence held back the development of microbiology. In the 1850s and 1860s, yeasts were established as microbes and responsible for alcoholic fermentation, and this led to the study of the rôle of bacteria in lactic and other fermentations, as well as bacterial pathogenicity. At this time, there were difficulties in distinguishing between the activities of microbes and of extracellular enzymes. Between 1884 and 1894, Emil Fischer's study of sugar utilization by yeasts generated an understanding of enzymic specificity and the nature of enzyme-substrate complexes.

Diversity and antifungal resistance patterns of prevalent opportunistic pathogenic yeasts colonizing the oral cavities of asymptomatic human immunodeficiency virus-infected individuals, and their relation to CD4+ counts

PubMed Central

Kumar, Deepa Anil; Muralidhar, Sumathi; Banerjee, Uma; Basir, Seemi Farhat; Mathur, Purva; Khan, Luqman Ahmad

2015-01-01

Background: Yeasts are important opportunistic pathogens, in individuals infected with human immunodeficiency virus (HIV). Yeast species inhabiting the oral mucosa of HIV-infected persons can act as source of oral lesions, especially as the individual progresses towards immunocompromised state. Present study was conducted to evaluate the diversity of yeasts in oral cavities of asymptomatic HIV-infected persons and their association with CD4+ cell counts. Materials and Methods: 100 HIV seropositive subjects and 100 healthy controls were screened for oral yeast carriage using standard procedures. Results: Of the 100 HIV-seropositive persons screened, 48 were colonized by different yeasts, either alone or in association with another species. Candida albicans was the most common species (56.90%) while non C. albicans Candida (NCAC) accounted for 39.65%. Among NCAC, Candida tropicalis and Candida krusei were most common. One isolate each of rare opportunistic pathogenic yeasts, Geotrichum candidum and Saccharomyces cereviseae, was recovered. The control group had an oral candidal carriage rate of 23%; C. albicans was the predominant species, followed by Candida glabrata, C. tropicalis and Candida parapsilosis. Antifungal susceptibility testing revealed no resistance in C. albicans, to the commonly used antifungal agents, whereas resistance or dose dependent susceptibility to fluconazole was observed in some of the NCAC species. Conclusion: Oral carriage of opportunistic pathogenic yeasts was greater in HIV-seropositive persons heading towards immunocompromised state, as evidenced by their CD4+ cell count. The predominant yeast isolated in this study (C. albicans), was found to be susceptible to commonly used antifungals. PMID:26392655

Evaluation of damage induced by Kwkt and Pikt zymocins against Brettanomyces/Dekkera spoilage yeast, as compared to sulphur dioxide.

PubMed

Oro, L; Ciani, M; Bizzaro, D; Comitini, F

2016-07-01

Over the last few decades, the use of zymocins as biological tools to counteract contamination by spoilage yeast in beverages and food has been widely studied. This study examined the damage induced by the Kwkt and Pikt, two zymocins produced by Kluyeromyces wickerhamii and Wickerhanomyces anomalus, respectively, with antimicrobial activity against Brettanomyces/Dekkera wine-spoilage yeast. The physiological and biochemical characterization of both of these proteins revealed that only Pikt showed a strict relationship between β-glucosidase activity and killer activity. The minimum inhibitory concentrations and minimum fungicidal concentrations of Kwkt and Pikt showed inhibitory activities against Brettanomyces/Dekkera yeast. Cytofluorimetric evaluation of cell death was based on both cell membrane permeability and cell metabolism, using fluorescence techniques under increasing zymocin levels over different incubation times. The antimicrobial actions of Kwkt and Pikt were also compared with the mode of action of sulphur dioxide. In this last case, the induction of the viable but noncultivable (VBNC) state was confirmed, with the consequent recovery of Brettanomyces yeast after medium replacement. In contrast, Kwkt and Pikt caused irreversible death of these yeast, without recovery of sensitive cells. Kwkt and Pikt could be proposed as fungistatic or fungicide biocontrol agents in winemaking to control the colonization and development of Brettanomyces/Dekkera yeasts. These data support the potential use of zymocins to reduce wine contamination as an alternative to sulphur dioxide that act on sensitive cells. Differently from sulphur dioxide, that could induce a reversible VBNC state, Kwkt and Pikt determine the irreversible damage on sensitive yeasts, ensuring the complete control of spoilage Brettanomyces yeast. © 2016 The Society for Applied Microbiology.

Post-fermentative production of glutathione by baker's yeast (S. cerevisiae) in compressed and dried forms.

PubMed

Musatti, Alida; Manzoni, Matilde; Rollini, Manuela

2013-01-25

The study was aimed at investigating the best biotransformation conditions to increase intracellular glutathione (GSH) levels in samples of baker's yeast (Saccharomyces cerevisiae) employing either the commercially available compressed and dried forms. Glucose, GSH precursors amino acids, as well as other cofactors, were dissolved in a biotransformation solution and yeast cells were added (5%dcw). Two response surface central composite designs (RSCCDs) were performed in sequence: in the first step the influence of amino acid composition (cysteine, glycine, glutamic acid and serine) on GSH accumulation was investigated; once their formulation was set up, the influence of other components was studied. Initial GSH content was found 0.53 and 0.47%dcw for compressed and dried forms. GSH accumulation ability of baker's yeast in compressed form was higher at the beginning of shelf life, that is, in the first week, and a maximum of 2.04%dcw was obtained. Performance of yeast in dried form was not found satisfactory, as the maximum GSH level was 1.18%dcw. When cysteine lacks from the reaction solution, yeast cells do not accumulate GSH. With dried yeast, the highest GSH yields occurred when cysteine was set at 3 g/L, glycine and glutamic acid at least at 4 g/L, without serine. Employing compressed yeast, the highest GSH yields occurred when cysteine and glutamic acid were set at 2-3 g/L, while glycine and serine higher than 2 g/L. Results allowed to set up an optimal and feasible procedure to obtain GSH-enriched yeast biomass, with up to threefold increase with respect to initial content. Copyright © 2012 Elsevier B.V. All rights reserved.

[A study on the bond interface between low-fusing dental porcelain and pure titanium].

PubMed

Mo, A; Cen, Y; Liao, Y; Wang, J; Shi, X

2001-09-01

To evaluate the bond interface between low fusing dental porcelain and pure titanium by observing the topography and detecting the ionic diffusion in the interface area. The low fusing-porcelain La-porcelain produced by the authors or Vita Titankeramik porcelain was fused to the surfaces of pure titanium. The topography of the interface between pure titanium and porcelain, and the structure of experimental materials were observed with SEM. The state of ionic diffusion in the interface area was investigated with EPMA. Excellent permeation and diffusion of La-porcelain were observed on the surfaces of pure titanium. The diffusion of ions of stannum and silicon was discovered in the interface area. The microstructure of La-porcelain to pure titanium bond interface was finer than that of Vita Titankeramik porcelain. Excellent bond can be produced in the interface between La-porcelain and pure titanium. The bonding mechanism may involve mechanical bond and chemical bond. The ionic diffusion of stannum plays an important role in the bonding of porcelain to pure titanium.

Deuterium Abundance Toward WD2211-495: Results from the Far Ultraviolet Spectroscopic Explorer (FUSE) Mission

NASA Technical Reports Server (NTRS)

Hebrard, G.; Lemoine, M.; Vidal-Madjar, A.; Desert, J. M.; LecavelierdesEtangs, A.; Ferlet, R.; Wood, B. E.; Linsky, J. L.; Kruk, J. W.; Chayer, P.;

Improvement of fermentation ability under baking-associated stress conditions by altering the POG1 gene expression in baker's yeast.

PubMed

Sasano, Yu; Haitani, Yutaka; Hashida, Keisuke; Oshiro, Satoshi; Shima, Jun; Takagi, Hiroshi

2013-08-01

During the bread-making process, yeast cells are exposed to many types of baking-associated stress. There is thus a demand within the baking industry for yeast strains with high fermentation abilities under these stress conditions. The POG1 gene, encoding a putative transcription factor involved in cell cycle regulation, is a multicopy suppressor of the yeast Saccharomyces cerevisiae E3 ubiquitin ligase Rsp5 mutant. The pog1 mutant is sensitive to various stresses. Our results suggested that the POG1 gene is involved in stress tolerance in yeast cells. In this study, we showed that overexpression of the POG1 gene in baker's yeast conferred increased fermentation ability in high-sucrose-containing dough, which is used for sweet dough baking. Furthermore, deletion of the POG1 gene drastically increased the fermentation ability in bread dough after freeze-thaw stress, which would be a useful characteristic for frozen dough baking. Thus, the engineering of yeast strains to control the POG1 gene expression level would be a novel method for molecular breeding of baker's yeast. Copyright © 2013 Elsevier B.V. All rights reserved.

Newly generated interspecific wine yeast hybrids introduce flavour and aroma diversity to wines.

PubMed

Bellon, Jennifer R; Eglinton, Jeffery M; Siebert, Tracey E; Pollnitz, Alan P; Rose, Louisa; de Barros Lopes, Miguel; Chambers, Paul J

2011-08-01

Increasingly, winemakers are looking for ways to introduce aroma and flavour diversity to their wines as a means of improving style and increasing product differentiation. While currently available commercial yeast strains produce consistently sound fermentations, there are indications that sensory complexity and improved palate structure are obtained when other species of yeast are active during fermentation. In this study, we explore a strategy to increase the impact of non-Saccharomyces cerevisiae inputs without the risks associated with spontaneous fermentations, through generating interspecific hybrids between a S. cerevisiae wine strain and a second species. For our experiments, we used rare mating to produce hybrids between S. cerevisiae and other closely related yeast of the Saccharomyces sensu stricto complex. These hybrid yeast strains display desirable properties of both parents and produce wines with concentrations of aromatic fermentation products that are different to what is found in wine made using the commercial wine yeast parent. Our results demonstrate, for the first time, that the introduction of genetic material from a non-S. cerevisiae parent into a wine yeast background can impact favourably on the wine flavour and aroma profile of a commercial S. cerevisiae wine yeast.

The Genetics of Non-conventional Wine Yeasts: Current Knowledge and Future Challenges.

PubMed

Masneuf-Pomarede, Isabelle; Bely, Marina; Marullo, Philippe; Albertin, Warren

2015-01-01

Saccharomyces cerevisiae is by far the most widely used yeast in oenology. However, during the last decade, several other yeasts species has been purposed for winemaking as they could positively impact wine quality. Some of these non-conventional yeasts (Torulaspora delbrueckii, Metschnikowia pulcherrima, Pichia kluyveri, Lachancea thermotolerans, etc.) are now proposed as starters culture for winemakers in mixed fermentation with S. cerevisiae, and several others are the subject of various studies (Hanseniaspora uvarum, Starmerella bacillaris, etc.). Along with their biotechnological use, the knowledge of these non-conventional yeasts greatly increased these last 10 years. The aim of this review is to describe the last updates and the current state-of-art of the genetics of non-conventional yeasts (including S. uvarum, T. delbrueckii, S. bacillaris, etc.). We describe how genomics and genetics tools provide new data into the population structure and biodiversity of non-conventional yeasts in winemaking environments. Future challenges will lie on the development of selection programs and/or genetic improvement of these non-conventional species. We discuss how genetics, genomics and the advances in next-generation sequencing will help the wine industry to develop the biotechnological use of non-conventional yeasts to improve the quality and differentiation of wines.

In vitro studies on the translocation of acid phosphatase into the endoplasmic reticulum of the yeast Saccharomyces cerevisiae.

PubMed

Krebs, H O; Hoffschulte, H K; Müller, M

1989-05-01

We demonstrate here the in vitro translocation of yeast acid phosphatase into rough endoplasmic reticulum. The precursor of the repressible acid phosphatase from Saccharomyces cerevisiae encoded by the PHO5 gene, was synthesized in a yeast lysate programmed with in vitro transcribed PHO5 mRNA. In the presence of yeast rough microsomes up to 16% of the acid phosphatase synthesized was found to be translocated into the microsomes, as judged by proteinase resistance, and fully core-glycosylated. The translocation efficiency however, decreased to 3% if yeast rough microsomes were added after synthesis of acid phosphatase had been terminated. When a wheat-germ extract was used for in vitro synthesis, the precursor of acid phosphatase was translocated into canine pancreatic rough microsomes and thereby core-glycosylated in a signal-recognition-particle-dependent manner. Replacing canine with yeast rough microsomes in the wheat-germ translation system, however, resulted in a significant decrease in the ability to translocate and glycosylate the precursor. Translocation and glycosylation were partially restored by a high-salt extract prepared from yeast ribosomes. The results presented here suggest that yeast-specific factors are needed to translocate and glycosylate acid phosphatase efficiently in vitro.

AskFuse Origins: System Barriers to Providing the Research That Public Health Practice and Policy Partners Say They Need

ERIC Educational Resources Information Center

Rushmer, Rosemary; Shucksmith, Janet

2018-01-01

In this paper the development of askFuse is used as a case study to illustrate contextual and system barriers to universities providing useful, usable and timely research evidence in response to local practice and policy partners' stated public health research needs. Entrenched systems (research excellence framework, academic career pathways,…

Interactions between yeast lees and wine polyphenols during simulation of wine aging: I. Analysis of remnant polyphenolic compounds in the resulting wines.

PubMed

Mazauric, Jean-Paul; Salmon, Jean-Michel

2005-07-13

Wine aging on yeast lees is a traditional enological practice used during the manufacture of wines. This technique has increased in popularity in recent years for the aging of red wines. Although wine polyphenols interact with yeast lees to a limited extent, such interactions have a large effect on the reactivity toward oxygen of wine polyphenolic compounds and yeast lees. Various domains of the yeast cell wall are protected by wine polyphenols from the action of extracellular hydrolytic enzymatic activities. Polysaccharides released during autolysis are thought to exert a significant effect on the sensory qualities of wine. We studied the chemical composition of polyphenolic compounds remaining in solution or adsorbed on yeast lees after various contact times during the simulation of wine aging. The analysis of the remnant polyphenols in the wine indicated that wine polyphenols adsorption on yeast lees follows biphasic kinetics. An initial and rapid fixation is followed by a slow, constant, and saturating fixation that reaches its maximum after about 1 week. Only very few monomeric phenolic compounds remained adsorbed on yeast lees, and no preferential adsorption of low or high polymeric size tannins occurred. The remnant condensed tannins in the wine contained fewer epigallocatechin units than the initial tannins, indicating that polar condensed tannins were preferentially adsorbed on yeast lees. Conversely, the efficiency of anthocyanin adsorption on yeast lees was unrelated to its polarity.

Distribution of tannin-'tolerant yeasts isolated from Miang, a traditional fermented tea leaf (Camellia sinensis var. assamica) in northern Thailand.

PubMed

Kanpiengjai, Apinun; Chui-Chai, Naradorn; Chaikaew, Siriporn; Khanongnuch, Chartchai

2016-12-05

Miang is a fermented food product prepared from the tea leaves of Camellia sinensis var. assamica, and is traditionally produced in mountainous areas of northern Thailand. Although Miang has a long history and reveals deep-rooted cultural involvement with local people in northern Thailand, little is known regarding its microbial diversity. Yeasts were isolated from 47 Miang samples collected from 28 sampling sites, including eight provinces in upper northern Thailand. A hundred and seven yeast isolates were recovered and identified within 14 species based on the comparison of the D1/D2 sequence of the large subunit (LSU) rRNA gene. Candida ethanolica was determined to be the dominant species that was frequently found in Miang together with minor resident yeast species. All yeast isolates demonstrated their tannin-tolerant capability when cultivated on yeast malt agar (YMA) containing 50g/l tannin, but nine isolates displayed clear zones forming around their colonies, e.g., Debaryomyces hansenii, Cyberlindnera rhodanensis, and Sporidiobolus ruineniae. The results obtained from a visual reading method of tannase revealed that all yeast isolates were positive for methyl gallate, indicating that they possess tannase activity. It is assumed that a tannin-tolerant ability is one of the most important factors for developing a yeast community in Miang. This research study is the first report to describe tannin-tolerant yeasts and yeast communities in traditionally fermented tea leaves. Copyright © 2016. Published by Elsevier B.V.

Influence of different floor management strategies of the vineyard on the natural yeast population associated with grape berries.

PubMed

Cordero-Bueso, Gustavo; Arroyo, Teresa; Serrano, Ana; Valero, Eva

2011-07-15

Some oenological practices, such as the massive utilisation of commercial yeast and the consequent colonisation of wineries, can contribute to reducing the native yeast biodiversity. In this context, the vineyard could be a reservoir of autochthonous yeasts of oenological interest. Thus, the evaluation of the influence of different agricultural parameters on the biodiversity of yeast population in the vineyard is necessary. This work shows the results of the influence of some floor management strategies of the vineyard in the natural yeast population associated with the grape-berries. With this objective, a three year sampling plan was designed in the Shiraz vineyards of the Madrid region using three floor management strategies: bare soil by tillage, bare soil maintained with herbicides and soil maintained with cover crop. The results of this study have shown that bare soil by tillage could be a sustainable alternative for managing the soil, due to the reduced use of agrochemicals and the resulting high yeasts biodiversity. Nevertheless, the presence of herbicides in the vineyard has a minor impact on the diversity of grape associated yeast communities, and this could have increased the yeast populations. Hence, from the fermentative yeasts' (like Saccharomyces) point of view, in hot and arid environments where soils may be affected by the tillage management, the best option could be the maintenance of the bare soil with the use of herbicides. Copyright © 2011 Elsevier B.V. All rights reserved.

Bulk damage and absorption in fused silica due to high-power laser applications

NASA Astrophysics Data System (ADS)

Nürnberg, F.; Kühn, B.; Langner, A.; Altwein, M.; Schötz, G.; Takke, R.; Thomas, S.; Vydra, J.

2015-11-01

Laser fusion projects are heading for IR optics with high broadband transmission, high shock and temperature resistance, long laser durability, and best purity. For this application, fused silica is an excellent choice. The energy density threshold on IR laser optics is mainly influenced by the purity and homogeneity of the fused silica. The absorption behavior regarding the hydroxyl content was studied for various synthetic fused silica grades. The main absorption influenced by OH vibrational excitation leads to different IR attenuations for OH-rich and low-OH fused silica. Industrial laser systems aim for the maximum energy extraction possible. Heraeus Quarzglas developed an Yb-doped fused silica fiber to support this growing market. But the performance of laser welding and cutting systems is fundamentally limited by beam quality and stability of focus. Since absorption in the optical components of optical systems has a detrimental effect on the laser focus shift, the beam energy loss and the resulting heating has to be minimized both in the bulk materials and at the coated surfaces. In collaboration with a laser research institute, an optical finisher and end users, photo thermal absorption measurements on coated samples of different fused silica grades were performed to investigate the influence of basic material properties on the absorption level. High purity, synthetic fused silica is as well the material of choice for optical components designed for DUV applications (wavelength range 160 nm - 260 nm). For higher light intensities, e.g. provided by Excimer lasers, UV photons may generate defect centers that effect the optical properties during usage, resulting in an aging of the optical components (UV radiation damage). Powerful Excimer lasers require optical materials that can withstand photon energy close to the band gap and the high intensity of the short pulse length. The UV transmission loss is restricted to the DUV wavelength range below 300 nm and consists of three different absorption bands centered at 165 nm (peroxy radicals), 215 nm (E'-center), and 265 nm (non-bridging oxygen hole center (NBOH)), which change the transmission behavior of material.

A survey of yeast from the Yarrowia clade for lipid production in dilute-acid pretreated lignocellulosic biomass hydrolysate

USDA-ARS?s Scientific Manuscript database

Yarrowia lipolytica is an oleaginous yeast species that has attracted attention as a model organism for synthesis of single cell oil. Among over 50 isolates of Y. lipolytica identified, only a few of the strains have been studied extensively. Furthermore, 12 other yeast species were recently assigne...

30 CFR 28.2 - Approved fuses.

Code of Federal Regulations, 2010 CFR

2010-07-01

... MINING PRODUCTS FUSES FOR USE WITH DIRECT CURRENT IN PROVIDING SHORT-CIRCUIT PROTECTION FOR TRAILING...-circuit protection for trailing cables in coal mines only where such fuses are: (1) The same in all...

Biodeterioration of medical-grade silicone rubber used for voice prostheses: a SEM study.

PubMed

Neu, T R; Van der Mei, H C; Busscher, H J; Dijk, F; Verkerke, G J

1993-05-01

Silicone voice prostheses used for rehabilitation of speech after total laryngectomy are inserted in an non-sterile habitat. Deposits on explanted Groningen Button voice prostheses revealed a biofilm, due to heavy colonization of the silicone surface by bacteria and yeasts. Furthermore, it was demonstrated by scanning electron microscopy on sectioned explants that the silicone material was deteriorated by filamentous and vegetative yeast cells. The different explants showed a variety of sharp-edged, discrete yeast colonies. The yeasts grew just under the silicone surface and up to 700 microns into the silicone material. Finally, nine different types of defects in the silicone material created by the yeasts are described. This deterioration of the silicone by yeasts seems to be the main reason for the failure and the frequent replacement of the prostheses. The mechanisms of silicone deterioration are still hypothetical.

The contribution of glutathione to the destabilizing effect of yeast on wheat dough.

PubMed

Verheyen, C; Albrecht, A; Herrmann, J; Strobl, M; Jekle, M; Becker, T

2015-04-15

Any factor which impairs the development of the gluten network affects the gas retention capacity and the overall baking performance. This study aimed to examine why rising yeast concentrations (Saccharomyces cerevisiae) decrease the dough elasticity in an asymptotic manner. Since in 27 commercial fresh and dry yeasts up to 81 mg glutathione (GSH) per 1g dry sample were found. Through the addition of reduced GSH in dough without yeast, the extent of dough weakening was analysed. Indeed rheological measurements confirmed that yeast-equivalent levels of GSH had a softening effect and during 3h fermentation the weakening coefficient increased from 0.3% to 20.4% in a Rheofermentometer. The present results indicate that free -SH compounds, as represented by GSH, considerably contribute to the softening of dough through dead yeast cells. Copyright © 2014 Elsevier Ltd. All rights reserved.

Tolerance of budding yeast Saccharomyces cerevisiae to ultra high pressure

NASA Astrophysics Data System (ADS)

Shibata, M.; Torigoe, M.; Matsumoto, Y.; Yamamoto, M.; Takizawa, N.; Hada, Y.; Mori, Y.; Takarabe, K.; Ono, F.

2014-05-01

Our studies on the tolerance of plants and animals against very high pressure of several GPa have been extended to a smaller sized fungus, the budding yeast Saccharomyces cerevisiae. Several pieces of budding yeast (dry yeast) were sealed in a small teflon capsule with a liquid pressure medium fluorinate, and exposed to 7.5 GPa by using a cubic anvil press. The pressure was kept constant for various duration of time from 2 to 24 h. After the pressure was released, the specimens were brought out from the teflon capsule, and they were cultivated on a potato dextrose agar. It was found that the budding yeast exposed to 7.5 GPa for up to 6 h showed multiplication. However, those exposed to 7.5 GPa for longer than 12 h were found dead. The high pressure tolerance of budding yeast is a little weaker than that of tardigrades.

Yeast Methylotrophy and Autophagy in a Methanol-Oscillating Environment on Growing Arabidopsis thaliana Leaves

PubMed Central

Kawaguchi, Kosuke; Yurimoto, Hiroya; Oku, Masahide; Sakai, Yasuyoshi

2011-01-01

The yeast Candida boidinii capable of growth on methanol proliferates and survives on the leaves of Arabidopsis thaliana. The local methanol concentration at the phyllosphere of growing A. thaliana exhibited daily periodicity, and yeast cells responded by altering both the expression of methanol-inducible genes and peroxisome proliferation. Even under these dynamically changing environmental conditions, yeast cells proliferated 3 to 4 times in 11 days. Among the C1-metabolic enzymes, enzymes in the methanol assimilation pathway, but not formaldehyde dissimilation or anti-oxidizing enzymes, were necessary for yeast proliferation at the phyllosphere. Furthermore, both peroxisome assembly and pexophagy, a selective autophagy pathway that degrades peroxisomes, were necessary for phyllospheric proliferation. Thus, the present study sheds light on the life cycle and physiology of yeast in the natural environment at both the molecular and cellular levels. PMID:21966472

Matrilin-1 expression is increased in the vertebral column of Atlantic salmon (Salmo salar L.) individuals displaying spinal fusions.

PubMed

Pedersen, Mona E; Takle, Harald; Ytteborg, Elisabeth; Veiseth-Kent, Eva; Enersen, Grethe; Færgestad, Ellen; Baeverfjord, Grete; Hannesson, Kirsten O

2011-12-01

We have previously characterized the development of vertebral fusions induced by elevated water temperature in Atlantic salmon. Molecular markers of bone and cartilage development together with histology were used to understand the complex pathology and mechanism in the development of this spinal malformation. In this study, we wanted to use proteomics, a non-hypothetical approach to screen for possible new markers involved in the fusion process. Proteins extracted from non-deformed and fused vertebrae of Atlantic salmon were therefore compared by two-dimensional electrophoresis (2DE) and MALDI-TOF analysis. Data analysis of protein spots in the 2DE gels demonstrated matrilin-1, also named cartilage matrix protein, to be the most highly up-regulated protein in fused compared with non-deformed vertebrae. Furthermore, real-time PCR analysis showed strong up-regulation of matrilin-1 mRNA in fused vertebrae. Immunohistochemistry demonstrated induced matrilin-1 expression in trans-differentiating cells undergoing a metaplastic shift toward chondrocytes in fusing vertebrae, whereas abundant expression was demonstrated in cartilaginous tissue and chordocytes of both non-deformed and fused vertebrae. These results identifies matrilin-1 as a new interesting candidate in the fusion process, and ratify the use of proteomic as a valuable technique to screen for markers involved in vertebral pathogenesis.

Co-fuse: a new class discovery analysis tool to identify and prioritize recurrent fusion genes from RNA-sequencing data.

PubMed

Paisitkriangkrai, Sakrapee; Quek, Kelly; Nievergall, Eva; Jabbour, Anissa; Zannettino, Andrew; Kok, Chung Hoow

2018-06-07

Recurrent oncogenic fusion genes play a critical role in the development of various cancers and diseases and provide, in some cases, excellent therapeutic targets. To date, analysis tools that can identify and compare recurrent fusion genes across multiple samples have not been available to researchers. To address this deficiency, we developed Co-occurrence Fusion (Co-fuse), a new and easy to use software tool that enables biologists to merge RNA-seq information, allowing them to identify recurrent fusion genes, without the need for exhaustive data processing. Notably, Co-fuse is based on pattern mining and statistical analysis which enables the identification of hidden patterns of recurrent fusion genes. In this report, we show that Co-fuse can be used to identify 2 distinct groups within a set of 49 leukemic cell lines based on their recurrent fusion genes: a multiple myeloma (MM) samples-enriched cluster and an acute myeloid leukemia (AML) samples-enriched cluster. Our experimental results further demonstrate that Co-fuse can identify known driver fusion genes (e.g., IGH-MYC, IGH-WHSC1) in MM, when compared to AML samples, indicating the potential of Co-fuse to aid the discovery of yet unknown driver fusion genes through cohort comparisons. Additionally, using a 272 primary glioma sample RNA-seq dataset, Co-fuse was able to validate recurrent fusion genes, further demonstrating the power of this analysis tool to identify recurrent fusion genes. Taken together, Co-fuse is a powerful new analysis tool that can be readily applied to large RNA-seq datasets, and may lead to the discovery of new disease subgroups and potentially new driver genes, for which, targeted therapies could be developed. The Co-fuse R source code is publicly available at https://github.com/sakrapee/co-fuse .

PMAA-stabilized ferrofluid/chitosan/yeast composite for bioapplications

NASA Astrophysics Data System (ADS)

Baldikova, Eva; Prochazkova, Jitka; Stepanek, Miroslav; Hajduova, Jana; Pospiskova, Kristyna; Safarikova, Mirka; Safarik, Ivo

2017-04-01

A simple, one-pot process for the preparation of magnetically responsive yeast-based biocatalysts was developed. Saccharomyces cerevisiae, Candida utilis and Kluyveromyces lactis cells were successfully incorporated into chitosan gel magnetically modified with poly(methacrylic acid)-stabilized magnetic fluid (PMAA-FF) during its formation. Magnetic PMAA-FF/chitosan/yeast composites were efficiently employed for invert sugar production. The dependence of invertase activity on used yeast, amount of magnetic biocatalyst, agitation time and after reuse was studied in detail. The tested magnetic biocatalysts retained at least 69% of their initial activity after 8 reuse cycles.

[Antivirus effect of polysaccharides of brewer yeast in vitro].

PubMed

Li, F; Shi, Y; Guan, X; Zhang, S; Tian, T

1998-03-01

The antivirus effect of polysaccharides of brewer yeast from yeast mud on 13 kinds of viruses including DNA and RNA virus along with their mechanisms were studied. The result showed that this effect was remarkable on the infections with poliovirus III, adenovirus III, ECHO6 virus, enterovirus 71, vesicular stomatitis virus, herpesvirus I, II, coxsackie A16 virus and coxsackie B3 virus. The polysaccharides of brewer yeast could also inhibit the development of cytopathic effect(CPE) and protect cultural cells from being infected with the above viruses.

Evolutionary biology through the lens of budding yeast comparative genomics.

PubMed

Marsit, Souhir; Leducq, Jean-Baptiste; Durand, Éléonore; Marchant, Axelle; Filteau, Marie; Landry, Christian R

2017-10-01

The budding yeast Saccharomyces cerevisiae is a highly advanced model system for studying genetics, cell biology and systems biology. Over the past decade, the application of high-throughput sequencing technologies to this species has contributed to this yeast also becoming an important model for evolutionary genomics. Indeed, comparative genomic analyses of laboratory, wild and domesticated yeast populations are providing unprecedented detail about many of the processes that govern evolution, including long-term processes, such as reproductive isolation and speciation, and short-term processes, such as adaptation to natural and domestication-related environments.

Developmental Toxicity Studies with Pregabalin in Rats: Significance of Alterations in Skull Bone Morphology.

PubMed

Morse, Dennis C; Henck, Judith W; Bailey, Steven A

2016-04-01

Pregabalin was administered to pregnant Wistar rats during organogenesis to evaluate potential developmental toxicity. In an embryo-fetal development study, compared with controls, fetuses from pregabalin-treated rats exhibited increased incidence of jugal fused to maxilla (pregabalin 1250 and 2500 mg/kg) and fusion of the nasal sutures (pregabalin 2500 mg/kg). The alterations in skull development occurred in the presence of maternal toxicity (reduced body weight gain) and developmental toxicity (reduced fetal body weight and increased skeletal variations), and were initially classified as malformations. Subsequent investigative studies in pregnant rats treated with pregabalin during organogenesis confirmed the advanced jugal fused to maxilla, and fusion of the nasal sutures at cesarean section (gestation day/postmating day [PMD] 21) in pregabalin-treated groups. In a study designed to evaluate progression of skull development, advanced jugal fused to maxilla and fusion of the nasal sutures was observed on PMD 20-25 and PMD 21-23, respectively (birth occurs approximately on PMD 22). On postnatal day (PND) 21, complete jugal fused to maxilla was observed in the majority of control and 2500 mg/kg offspring. No treatment-related differences in the incidence of skull bone fusions occurred on PND 21, indicating no permanent adverse outcome. Based on the results of the investigative studies, and a review of historical data and scientific literature, the advanced skull bone fusions were reclassified as anatomic variations. Pregabalin was not teratogenic in rats under the conditions of these studies. © 2016 Wiley Periodicals, Inc.

KSC-99pp0494

NASA Image and Video Library

1999-05-03

Workers at Hangar AE, Cape Canaveral Air Station, maneuver an overhead crane toward NASA's Far Ultraviolet Spectroscopic Explorer (FUSE) satellite standing between vertical workstands. The crane will lift FUSE to move it onto the Payload Attach Fitting (PAF) in front of it. FUSE is undergoing a functional test of its systems, plus installation of flight batteries and solar arrays. Developed by The Johns Hopkins University under contract to Goddard Space Flight Center, Greenbelt, Md., FUSE will investigate the origin and evolution of the lightest elements in the universe hydrogen and deuterium. In addition, the FUSE satellite will examine the forces and process involved in the evolution of the galaxies, stars and planetary systems by investigating light in the far ultraviolet portion of the electromagnetic spectrum. FUSE is scheduled to be launched May 27 aboard a Boeing Delta II rocket at Launch Complex 17

The Effect of Drycleaning Moisture on Fused Cloth Systems

DTIC Science & Technology

1989-03-01

TECHNICAL REPORT NATICK/TR-89/024 et, THE EFFECT OF DRYCLEANING MOISTURE ON FUSED CLOTH SYSTEMS BY ELIZABETH J. MORELAND International...MOISTUP.E ON FUSED CLOTH SYSTEMS 12. PERSONAL AUTMOR(S) Elizabeth J. MorelanJ 13«. TYPE OF REPORT Final Technical Report 13b. TIME COVERED...This project was initiated to investigate the effect of moisture in drycleaning systems on preselected fused cloth structures. Adverse surface

Halting the fuse discharge propagation using optical fiber microwires.

PubMed

Rocha, A M; Fernandes, G; Domingues, F; Niehus, M; Pinto, A N; Facão, M; André, P S

2012-09-10

We report and analyze the halting of the fuse effect propagation in optical fiber microwires. The increase of the mode field diameter in the tapered region decreases the optical intensity resulting in the extinction of the fuse effect. This fiber element presents a low insertion loss and can be introduced in the optical network in order to protect the active equipment from the damage caused by the fuse effect.

Gradient-based multiresolution image fusion.

PubMed

Petrović, Valdimir S; Xydeas, Costas S

2004-02-01

A novel approach to multiresolution signal-level image fusion is presented for accurately transferring visual information from any number of input image signals, into a single fused image without loss of information or the introduction of distortion. The proposed system uses a "fuse-then-decompose" technique realized through a novel, fusion/decomposition system architecture. In particular, information fusion is performed on a multiresolution gradient map representation domain of image signal information. At each resolution, input images are represented as gradient maps and combined to produce new, fused gradient maps. Fused gradient map signals are processed, using gradient filters derived from high-pass quadrature mirror filters to yield a fused multiresolution pyramid representation. The fused output image is obtained by applying, on the fused pyramid, a reconstruction process that is analogous to that of conventional discrete wavelet transform. This new gradient fusion significantly reduces the amount of distortion artefacts and the loss of contrast information usually observed in fused images obtained from conventional multiresolution fusion schemes. This is because fusion in the gradient map domain significantly improves the reliability of the feature selection and information fusion processes. Fusion performance is evaluated through informal visual inspection and subjective psychometric preference tests, as well as objective fusion performance measurements. Results clearly demonstrate the superiority of this new approach when compared to conventional fusion systems.

Chromatin and Transcription in Yeast

PubMed Central

Rando, Oliver J.; Winston, Fred

2012-01-01

Understanding the mechanisms by which chromatin structure controls eukaryotic transcription has been an intense area of investigation for the past 25 years. Many of the key discoveries that created the foundation for this field came from studies of Saccharomyces cerevisiae, including the discovery of the role of chromatin in transcriptional silencing, as well as the discovery of chromatin-remodeling factors and histone modification activities. Since that time, studies in yeast have continued to contribute in leading ways. This review article summarizes the large body of yeast studies in this field. PMID:22345607