Physiological response of Pichia pastoris GS115 to methanol-induced high level production of the Hepatitis B surface antigen: catabolic adaptation, stress responses, and autophagic processes

PubMed Central

2012-01-01

Background Pichia pastoris is an established eukaryotic host for the production of recombinant proteins. Most often, protein production is under the control of the strong methanol-inducible aox1 promoter. However, detailed information about the physiological alterations in P. pastoris accompanying the shift from growth on glycerol to methanol-induced protein production under industrial relevant conditions is missing. Here, we provide an analysis of the physiological response of P. pastoris GS115 to methanol-induced high-level production of the Hepatitis B virus surface antigen (HBsAg). High product titers and the retention of the protein in the endoplasmic reticulum (ER) are supposedly of major impact on the host physiology. For a more detailed understanding of the cellular response to methanol-induced HBsAg production, the time-dependent changes in the yeast proteome and ultrastructural cell morphology were analyzed during the production process. Results The shift from growth on glycerol to growth and HBsAg production on methanol was accompanied by a drastic change in the yeast proteome. In particular, enzymes from the methanol dissimilation pathway started to dominate the proteome while enzymes from the methanol assimilation pathway, e.g. the transketolase DAS1, increased only moderately. The majority of methanol was metabolized via the energy generating dissimilatory pathway leading to a corresponding increase in mitochondrial size and numbers. The methanol-metabolism related generation of reactive oxygen species induced a pronounced oxidative stress response (e.g. strong increase of the peroxiredoxin PMP20). Moreover, the accumulation of HBsAg in the ER resulted in the induction of the unfolded protein response (e.g. strong increase of the ER-resident disulfide isomerase, PDI) and the ER associated degradation (ERAD) pathway (e.g. increase of two cytosolic chaperones and members of the AAA ATPase superfamily) indicating that potential degradation of HBsAg could

An improved Abbott ARCHITECT assay for the detection of hepatitis B virus surface antigen (HBsAg).

PubMed

Lou, Sheng C; Pearce, Sandra K; Lukaszewska, Teresa X; Taylor, Russell E; Williams, Gregg T; Leary, Thomas P

2011-05-01

The sensitive and accurate detection of hepatitis B virus surface antigen (HBsAg) is critical to the identification of infection and the prevention of transfusion transmitted disease. Improvement in HBsAg assay sensitivity is essential to reduce the window to detect an acute HBV infection. Additionally, the sensitive detection of HBsAg mutants that continue to evolve due to vaccine escape, immune selection and an error prone reverse transcriptase is a necessity. A fully automated HBsAg prototype assay on the Abbott ARCHITECT instrument was developed to improve sensitivity and mutant detection. This magnetic microparticle-based assay utilizes anti-HBsAg monoclonal antibodies to capture antigen present in serum or plasma. Captured antigen is then detected using anti-HBsAg antibody conjugated with the chemiluminescent compound, acridinium. The sensitivity of the ARCHITECT HBsAg prototype assay was improved as compared to the current ARCHITECT, PRISM, and competitor HBsAg assays. The enhancement in assay sensitivity was demonstrated by the use of commercially available HBV seroconversion panels. The prototype assay detected more panel members (185 of 383) vs. the current ARCHITECT (171), PRISM (181), or competitor HBsAg assays (73/140 vs. 62/140, respectively). The ARCHITECT prototype assay also efficiently detected all mutants evaluated. Finally, the sensitivity improvement did not compromise the specificity of the assay (99.94%). An improved Abbott ARCHITECT HBsAg prototype assay with enhanced detection of HBsAg and HBsAg mutants, as well as equivalent specificity was developed for the detection, diagnosis, and management of HBV infection. Copyright © 2011 Elsevier B.V. All rights reserved.

Glucose-Induced Acidification in Yeast Cultures

ERIC Educational Resources Information Center

Myers, Alan; Bourn, Julia; Pool, Brynne

2005-01-01

We present an investigation (for A-level biology students and equivalent) into the mechanism of glucose-induced extracellular acidification in unbuffered yeast suspensions. The investigation is designed to enhance understanding of aspects of the A-level curriculum that relate to the phenomenon (notably glucose catabolism) and to develop key skills…

A novel therapeutic hepatitis B vaccine induces cellular and humoral immune responses and breaks tolerance in hepatitis B virus (HBV) transgenic mice.

PubMed

Buchmann, Pascale; Dembek, Claudia; Kuklick, Larissa; Jäger, Clemens; Tedjokusumo, Raindy; von Freyend, Miriam John; Drebber, Uta; Janowicz, Zbigniew; Melber, Karl; Protzer, Ulrike

2013-02-06

Therapeutic vaccines are currently being developed for chronic hepatitis B and C. As an alternative to long-term antiviral treatment or to support only partially effective therapy, they should activate the patient's immune system effectively to fight and finally control the virus. A paradigm of therapeutic vaccination is the potent induction of T-cell responses against key viral antigens - besides activation of a humoral immune response. We have evaluated the potential of a novel vaccine formulation comprising particulate hepatitis B surface (HBsAg) and core antigen (HBcAg), and the saponin-based ISCOMATRIX™ adjuvant for its ability to stimulate T and B cell responses in C57BL/6 mice and its ability to break tolerance in syngeneic HBV transgenic (HBVtg) mice. In C57BL/6 mice, the vaccine induced multifunctional HBsAg- and HBcAg-specific CD8+ T cells detected by staining for IFNγ, TNFα and IL-2, as well as high antibody titers against both antigens. Vaccination of HBVtg animals induced potent HBsAg- and HBcAg-specific CD8+ T-cell responses in spleens and HBcAg-specific CD8+ T-cell responses in livers as well as anti-HBs seroconversion two weeks post injection. Vaccination further reduced HBcAg expression in livers of HBVtg mice without causing liver damage. In summary, this study demonstrates therapeutic efficacy of a novel vaccine formulation in a mouse model of immunotolerant, chronic HBV infection. Copyright © 2013 Elsevier Ltd. All rights reserved.

Seroclearance of Hbsag in Chronic Hepatitis B Virus Patients on Lamivudine Therapy: A 10 Year Experience.

PubMed

Sali, Shahnaz; Merza, Muayad A; Saadat, Sina; Mustafa, Nazik H; Queiky, Farzam; Yadegarynia, Davood

2015-04-02

The aim of this study was to determine hepatitis B surface antigen (HBsAg) seroclearance rate among patients treated with lamivudine at a specialized tertiary care referral hospital in Tehran, Iran. All patients on lamivudine (biovudin®) therapy at a dose of 100 mg/day, who showed seroclearnace between March 2001 and September 2011 were recruited. The main evaluation parameters were duration of HBsAg seroclearance and duration of HBsAg seroconversion. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were evaluated using standard methods. HBsAg seroclearance was defined as two consecutive negative serums HBsAg at least 6 months apart, whereas HBsAg seroconversion was defined as the disappearance of serum HBsAg and the presence of anti-HBs for >6 months. A total of 203 chronic HBV patients treated with lamivudine at a dose of 100 mg/day were included in the study. HBsAg seroclearance and seroconversion were observed in 11 patients after the initiation of the lamivudine therapy. Overall, in lamivudine responder patients, the mean time to HBsAg seroclearance was 26.90±10.93 months (range: 12-48 months). Furthermore, the responders showed seroconversion after a mean time of 26.90±11.08 months from the initiation of lamivudine therapy. When comparing the characteristics of those who have responded to lamivudine and those who have not responded, baseline HBV-DNA levels was significantly lower in responder than non responder patients (p<0.001). Meantime, there was no difference in age, sex, baseline ALT, AST and liver biopsy score between lamivudine responder and lamivudine non-responder patients. Despite introduction of tenofovir and entecavir as first line treatment for chronic HBV infection, lamivudine remains to be a low cost, safe and effective drug for HBsAg seroclearnace.

Reactivation of hepatitis B virus infection with persistently negative HBsAg on three HBsAg assays in a lymphoma patient undergoing chemotherapy.

PubMed

Cheung, Wing-I; Chan, Henry Lik-Yuen; Leung, Vincent King-Sun; Tse, Chi-Hang; Fung, Kitty; Lin, Shek-Ying; Wong, Ann; Wong, Vincent Wai-Sun; Chau, Tai-Nin

2010-02-01

In patients with occult hepatitis B virus (HBV) infection, acute exacerbation may occur when they become immunocompromised. Usually, these patients develop hepatitis B surface antigen (HBsAg) seroreversion during the flare. Here we report on a patient with occult HBV infection, who developed HBV exacerbation after chemotherapy for diffuse large B-cell lymphoma. The resurgence of HBV DNA preceded the elevation of liver enzymes for 20 weeks. Atypically, despite high viraemia, serological tests showed persistently negative HBsAg using three different sensitive HBsAg assays (i.e., Architect, Murex and AxSYM). On comparing the amino acid sequence of the index patient with the consensus sequence, five mutations were found at pre-S1, five at pre-S2 and twenty-three mutations at the S region. Six amino acid mutations were located in the 'a' determinant, including P120T, K122R, M133T, F134L, D144A and G145A. The mutants K122R, F134L and G145A in our patient have not been tested for their sensitivity to Architect and Murex assays by the previous investigators and might represent the escape mutants to these assays.

Apoptotic signals induce specific degradation of ribosomal RNA in yeast

PubMed Central

Mroczek, Seweryn; Kufel, Joanna

2008-01-01

Organisms exposed to reactive oxygen species, generated endogenously during respiration or by environmental conditions, undergo oxidative stress. Stress response can either repair the damage or activate one of the programmed cell death (PCD) mechanisms, for example apoptosis, and finally end in cell death. One striking characteristic, which accompanies apoptosis in both vertebrates and yeast, is a fragmentation of cellular DNA and mammalian apoptosis is often associated with degradation of different RNAs. We show that in yeast exposed to stimuli known to induce apoptosis, such as hydrogen peroxide, acetic acid, hyperosmotic stress and ageing, two large subunit ribosomal RNAs, 25S and 5.8S, became extensively degraded with accumulation of specific intermediates that differ slightly depending on cell death conditions. This process is most likely endonucleolytic, is correlated with stress response, and depends on the mitochondrial respiratory status: rRNA is less susceptible to degradation in respiring cells with functional defence against oxidative stress. In addition, RNA fragmentation is independent of two yeast apoptotic factors, metacaspase Yca1 and apoptosis-inducing factor Aif1, but it relies on the apoptotic chromatin condensation induced by histone H2B modifications. These data describe a novel phenotype for certain stress- and ageing-related PCD pathways in yeast. PMID:18385160

Tolerance of pentose utilising yeast to hydrogen peroxide-induced oxidative stress.

PubMed

Spencer, Jennifer; Phister, Trevor G; Smart, Katherine A; Greetham, Darren

2014-03-17

Bioethanol fermentations follow traditional beverage fermentations where the yeast is exposed to adverse conditions such as oxidative stress. Lignocellulosic bioethanol fermentations involve the conversion of pentose and hexose sugars into ethanol. Environmental stress conditions such as osmotic stress and ethanol stress may affect the fermentation performance; however, oxidative stress as a consequence of metabolic output can also occur. However, the effect of oxidative stress on yeast with pentose utilising capabilities has yet to be investigated. Assaying for the effect of hydrogen peroxide-induced oxidative stress on Candida, Pichia and Scheffersomyces spp. has demonstrated that these yeast tolerate hydrogen peroxide-induced oxidative stress in a manner consistent with that demonstrated by Saccharomyces cerevisiae. Pichia guillermondii appears to be more tolerant to hydrogen peroxide-induced oxidative stress when compared to Candida shehatae, Candida succiphila or Scheffersomyces stipitis. Sensitivity to hydrogen peroxide-induced oxidative stress increased in the presence of minimal media; however, addition of amino acids and nucleobases was observed to increase tolerance. In particular adenine increased tolerance and methionine reduced tolerance to hydrogen peroxide-induced oxidative stress.

Tolerance of pentose utilising yeast to hydrogen peroxide-induced oxidative stress

PubMed Central

2014-01-01

Background Bioethanol fermentations follow traditional beverage fermentations where the yeast is exposed to adverse conditions such as oxidative stress. Lignocellulosic bioethanol fermentations involve the conversion of pentose and hexose sugars into ethanol. Environmental stress conditions such as osmotic stress and ethanol stress may affect the fermentation performance; however, oxidative stress as a consequence of metabolic output can also occur. However, the effect of oxidative stress on yeast with pentose utilising capabilities has yet to be investigated. Results Assaying for the effect of hydrogen peroxide-induced oxidative stress on Candida, Pichia and Scheffersomyces spp. has demonstrated that these yeast tolerate hydrogen peroxide-induced oxidative stress in a manner consistent with that demonstrated by Saccharomyces cerevisiae. Pichia guillermondii appears to be more tolerant to hydrogen peroxide-induced oxidative stress when compared to Candida shehatae, Candida succiphila or Scheffersomyces stipitis. Conclusions Sensitivity to hydrogen peroxide-induced oxidative stress increased in the presence of minimal media; however, addition of amino acids and nucleobases was observed to increase tolerance. In particular adenine increased tolerance and methionine reduced tolerance to hydrogen peroxide-induced oxidative stress. PMID:24636079

Identification of salivary components that induce transition of hyphae to yeast in Candida albicans.

PubMed

Leito, Jelani T D; Ligtenberg, Antoon J M; Nazmi, Kamran; Veerman, Enno C I

2009-10-01

Candida albicans, the major human fungal pathogen, undergoes a reversible morphological transition from single yeast cells to pseudohyphae and hyphae filaments. The hyphae form is considered the most invasive form of the fungus. The purpose of this study is to investigate the effect of saliva on hyphae growth of C. albicans. Candida albicans hyphae were inoculated in Roswell Park Memorial Institute medium with whole saliva, parotid saliva or buffer mimicking the saliva ion composition, and cultured for 18 h at 37 degrees C under aerobic conditions with 5% CO(2). Whole saliva and parotid saliva induced transition to yeast growth, whereas the culture with buffer remained in the hyphae form. Parotid saliva was fractionated on a reverse-phase C8 column and each fraction was tested for inducing transition to yeast growth. By immunoblotting, the salivary component in the active fraction was identified as statherin, a phosphoprotein of 43 amino acids that has been implicated in remineralization of the teeth. Synthetically made statherin induced transition of hyphae to yeast. By deletion of five amino acids at the negatively charged N-terminal site (DpSpSEE), yeast-inducing activity and binding to C. albicans were increased. In conclusion, statherin induces transition to yeast of C. albicans hyphae and may thus contribute to the oral defense against candidiasis.

Seroprevalence of HDV infection in HBsAg positive population in Ismailia, Egypt.

PubMed

Gomaa, Nahed I M; Metwally, Lobna A; Nemr, Nader; Younis, Soha

2013-01-01

Hepatitis D virus (HDV) is a defective RNA virus that needs hepatitis B surface antigen (HBsAg) from HBV for transmission. HDV can lead to fulminant hepatitis and the progression of chronic liver damage in patients with chronic hepatitis B. The aim of this study was to determine the seroprevalence of HDV among HBsAg positive individuals in Ismailia, Egypt. Serum samples were collected from 170 HBsAg positive healthy individuals from Suez Canal University blood bank over a one year period. All of them were seeking blood donation and found to be HBsAg positive during viral hepatitis screening workups which is routinely done prior to donation. Serum samples were screened for IgG antibodies to hepatitis delta virus (HDV) using enzyme-linked immunosorbent assay (ELISA) method. Anti-HDV antibodies were detected in 8 (4.7%) individuals aged from 29-43 years. Liver function tests showed that serum ALT and AST levels were elevated in the HBsAglanti-HDV positive cases. It is concluded that the rate of HDV infection in Ismailia is high and further investigation is needed to validate the findings and raise awareness about the risk of dual HBV and HDV infection.

Longitudinal profiles of highly sensitive hepatitis B surface antigen levels: re-evaluation of HBsAg seroclearance.

PubMed

Seto, Wai-Kay; Tanaka, Yasuhito; Wong, Danny K-H; Shinkai, Noboru; Cheung, Ka-Shing; Liu, Kevin S-H; Fung, James; Lai, Ching-Lung; Yuen, Man-Fung

2016-05-01

Serologic profiles after hepatitis B surface antigen (HBsAg) seroclearance in chronic hepatitis B (CHB) have not been well-studied. We employed a highly sensitive HBsAg (hs-HBsAg) assay (lower detection limit 0.5 mIU/ml), 100 times more sensitive than conventional HBsAg measurements. CHB patients achieving HBsAg seroclearance defined by conventional assays were followed up for serum hs-HBsAg, HBV DNA and antibody to HBsAg (anti-HBs) levels at 0 months, 6-12 months and 3-5 years after HBsAg seroclearance. Factors associated with hs-HBsAg detectability were determined. One hundred and nine patients were recruited; 94 (86.2%) were followed up to years 3-5; and 25 patients (22.9%) were on nucleoside analogue therapy for a median duration of 6.0 (range 1.5-12.7) years before HBsAg seroclearance. Detectable hs-HBsAg was noted in 88 (80.7%), 60 (55.0%) and 20 (21.3%) patients at 0 months, 6-12 months and 3-5 years respectively. At years 3-5, genotype B patients, when compared to genotype C patients, had a higher anti-HBs positive rate (63.2% and 41.1% respectively, P = 0.036). Serum anti-HBs positivity, when compared to persistent anti-HBs negativity, was associated with a lower rate of hs-HBsAg detection (7.4% and 40% respectively, P < 0.001). Multivariate analysis showed anti-HBs negativity at years 3-5 to be independently associated with persistently positive hs-HBsAg (P = 0.007, odds ratio 7.1, 95% confidence interval 1.7-29.3). Serum hs-HBsAg could detect HBsAg presence in a substantial proportion of CHB after HBsAg seroclearance defined by conventional assays, especially among anti-HBs negative individuals. Serum hs-HBsAg could potentially assist differentiating HBsAg-negative CHB from individuals with only past HBV exposure without carrier state. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Radiation-induced mitotic and meiotic aneuploidy in the yeast Saccharomyces cerevisiae.

PubMed

Parry, J M; Sharp, D; Tippins, R S; Parry, E M

1979-06-01

A number of genetic systems are described which in yeast may be used to monitor the induction of chromosome aneuploidy during both mitotic and meiotic cell division. Using these systems we have been able to demonstrate the induction of both monosomic and trisomic cells in mitotically dividing cells and disomic spores in meiotically dividing cells after both UV light and X-ray exposure. The frequency of UV-light-induced monosomic colonies were reduced by post-treatment with photoreactivity light and both UV-light- and X-ray-induced monosomic colonies were reduced by liquid holding post-treatment under non-nutrient conditions. Both responses indicate an involvement of DNA-repair mechanisms in the removal of lesions which may lead to monosomy in yeast. This was further confirmed by the response of an excision-defective yeast strain which showed considerably increased sensitivity to the induction of monosomic colonies by UV-light treatment at low doses. Yeast cultures irradiated at different stages of growth showed variation in their responses to both UV-light and X-rays, cells at the exponential phase of growth show maximum sensitivity to the induction of monosomic colonies at low doses whereas stationary phase cultures showed maximum induction of monosomic colonies at high does. The frequencies of X-ray-induced chromosome aneuploidy during meiosis leading to the production of disomic spores was shown to be dependent upon the stage of meiosis at which the yeast cells were exposed to radiation. Cells which had proceeded beyond the DNA synthetic stage of meiosis were shown to produce disomic spores at considerably lower radiation doses than those cells which had only recently been inoculated into sporulation medium. The results obtained suggest that the yeast sustem may be suitable for the study of sensitivities of the various stages of meiotic cell division to the induction of chromosome aneuploidy after radiation exposure.

Protein aggregation induced during glass bead lysis of yeast

PubMed Central

Papanayotou, Irene; Sun, Beimeng; Roth, Amy F.; Davis, Nicholas G.

2013-01-01

Yeast cell lysates produced by mechanical glass bead disruption are widely used in a variety of applications, including for the analysis of native function, e.g. protein–protein interaction, enzyme assays and membrane fractionations. Below, we report a striking case of protein denaturation and aggregation that is induced by this lysis protocol. Most of this analysis focuses on the type 1 casein kinase Yck2, which normally tethers to the plasma membrane through C-terminal palmitoylation. Surprisingly, when cells are subjected to glass bead disruption, non-palmitoylated, cytosolic forms of the kinase denature and aggregate, while membrane-associated forms, whether attached through their native palmitoyl tethers or through a variety of artificial membrane-tethering sequences, are wholly protected from denaturation and aggregation. A wider look at the yeast proteome finds that, while the majority of proteins resist glass bead-induced aggregation, a significant subset does, in fact, succumb to such denaturation. Thus, yeast researchers should be aware of this potential artifact when embarking on biochemical analyses that employ glass bead lysates to look at native protein function. Finally, we demonstrate an experimental utility for glass bead-induced aggregation, using its fine discrimination of membrane-associated from non-associated Yck2 forms to discern fractional palmitoylation states of Yck2 mutants that are partially defective for palmitoylation. PMID:20641011

Phenolic derivatives from soy flour ethanol extract are potent in vitro quinone reductase (QR) inducing agents.

PubMed

Bolling, Bradley W; Parkin, Kirk L

2008-11-26

The fractionation of soy flour directed by a cellular bioassay for induction of phase 2 detoxification enzymes was used to identify quinone reductase (QR) inducing agents. A phospholipid-depleted, 80% methanol-partitioned isolate from a crude ethanol extract of soy flour was resolved using normal phase medium-pressure liquid chromatography (MPLC). Early eluting fractions were found to be the most potent QR inducing agents among the separated fractions. Fraction 2 was the most potent, doubling QR at <2 mug/mL. Further fractionation of this isolate led to the identification of several constituents. Fatty acids and sn-1 and sn-2 monoacylglycerols were identified, but were not highly potent QR inducers. Benzofuran-3-carbaldehyde, 4-hydroxybenzaldeyde, 4-ethoxybenzoic acid, 4-ethoxycinnamic acid, benzofuran-2-carboxylic ethyl ester, and ferulic acid ethyl ester (FAEE) were also identified as QR inducing constituents of this fraction. FAEE was the most potent of the identified constituents, doubling QR specific activity at 3.2 muM in the cellular bioassay.

Effects of amino acid substitutions in hepatitis B virus surface protein on virion secretion, antigenicity, HBsAg and viral DNA.

PubMed

Xiang, Kuan-Hui; Michailidis, Eleftherios; Ding, Hai; Peng, Ya-Qin; Su, Ming-Ze; Li, Yao; Liu, Xue-En; Dao Thi, Viet Loan; Wu, Xian-Fang; Schneider, William M; Rice, Charles M; Zhuang, Hui; Li, Tong

2017-02-01

As important virological markers, serum hepatitis B surface antigen (HBsAg) and hepatitis B virus (HBV) DNA levels show large fluctuations among chronic hepatitis B patients. The aim of this study was to reveal the potential impact and mechanisms of amino acid substitutions in small hepatitis B surface proteins (SHBs) on serum HBsAg and HBV DNA levels. Serum samples from 230 untreated chronic hepatitis B patients with genotype C HBV were analyzed in terms of HBV DNA levels, serological markers of HBV infection and SHBs sequences. In vitro functional analysis of the identified SHBs mutants was performed. Among 230 SHBs sequences, there were 39 (16.96%) sequences with no mutation detected (wild-type) and 191 (83.04%) with single or multiple mutations. SHBs consist of 226 amino acids, of which 104 (46.02%) had mutations in our study. Some mutations (e.g., sE2G, sL21S, sR24K, sT47A/K, sC69stop (sC69∗), sL95W, sL98V, and sG145R) negatively correlated with serum HBsAg levels. HBsAg and HBV DNA levels from this group of patients had a positive correlation (r=0.61, p<0.001). In vitro analysis showed that these mutations reduced extracellular HBsAg and HBV DNA levels by restricting virion secretion and antibody binding capacity. Virion secretion could be rescued for sE2G, sC69∗, and sG145R by co-expression of wild-type HBsAg. The serum HBsAg levels were lower in untreated CHB patients with novel SHBs mutations outside the major antigenic region than those without mutations. Underlying mechanisms include impairment of virion secretion and lower binding affinity to antibodies used for HBsAg measurements. The hepatitis B surface antigen (HBsAg) is a major viral protein of the hepatitis B virus (HBV) secreted into patient blood serum and its quantification value serves as an important marker for the evaluation of chronic HBV infection and antiviral response. We found a few new amino acid substitutions in HBsAg associated with lower serum HBsAg and HBV DNA levels. These

A mitochondria-dependent pathway mediates the apoptosis of GSE-induced yeast.

PubMed

Cao, Sishuo; Xu, Wentao; Zhang, Nan; Wang, Yan; Luo, YunBo; He, Xiaoyun; Huang, Kunlun

2012-01-01

Grapefruit seed extract (GSE), which has powerful anti-fungal activity, can induce apoptosis in S. cerevisiae. The yeast cells underwent apoptosis as determined by testing for apoptotic markers of DNA cleavage and typical chromatin condensation by Terminal Deoxynucleotidyl Transferase-mediated dUTP Nick End Labeling (TUNEL) and 4,6'-diaminidino-2-phenylindole (DAPI) staining and electron microscopy. The changes of ΔΨmt (mitochondrial transmembrane potential) and ROS (reactive oxygen species) indicated that the mitochondria took part in the apoptotic process. Changes in this process detected by metabonomics and proteomics revealed that the yeast cells tenaciously resisted adversity. Proteins related to redox, cellular structure, membrane, energy and DNA repair were significantly increased. In this study, the relative changes in the levels of proteins and metabolites showed the tenacious resistance of yeast cells. However, GSE induced apoptosis in the yeast cells by destruction of the mitochondrial 60 S ribosomal protein, L14-A, and prevented the conversion of pantothenic acid to coenzyme A (CoA). The relationship between the proteins and metabolites was analyzed by orthogonal projections to latent structures (OPLS). We found that the changes of the metabolites and the protein changes had relevant consistency.

A Mitochondria-Dependent Pathway Mediates the Apoptosis of GSE-Induced Yeast

PubMed Central

Cao, Sishuo; Xu, Wentao; Zhang, Nan; Wang, Yan; Luo, YunBo; He, Xiaoyun; Huang, Kunlun

2012-01-01

Grapefruit seed extract (GSE), which has powerful anti-fungal activity, can induce apoptosis in S. cerevisiae. The yeast cells underwent apoptosis as determined by testing for apoptotic markers of DNA cleavage and typical chromatin condensation by Terminal Deoxynucleotidyl Transferase–mediated dUTP Nick End Labeling (TUNEL) and 4,6′-diaminidino-2-phenylindole (DAPI) staining and electron microscopy. The changes of ΔΨmt (mitochondrial transmembrane potential) and ROS (reactive oxygen species) indicated that the mitochondria took part in the apoptotic process. Changes in this process detected by metabonomics and proteomics revealed that the yeast cells tenaciously resisted adversity. Proteins related to redox, cellular structure, membrane, energy and DNA repair were significantly increased. In this study, the relative changes in the levels of proteins and metabolites showed the tenacious resistance of yeast cells. However, GSE induced apoptosis in the yeast cells by destruction of the mitochondrial 60 S ribosomal protein, L14-A, and prevented the conversion of pantothenic acid to coenzyme A (CoA). The relationship between the proteins and metabolites was analyzed by orthogonal projections to latent structures (OPLS). We found that the changes of the metabolites and the protein changes had relevant consistency. PMID:22403727

Potent L-lactic acid assimilation of the fermentative and heterothallic haploid yeast Saccharomyces cerevisiae NAM34-4C.

PubMed

Tomitaka, Masataka; Taguchi, Hisataka; Matsuoka, Masayoshi; Morimura, Shigeru; Kida, Kenji; Akamatsu, Takashi

2014-01-01

We screened an industrial thermotolerant Saccharomyces cerevisiae strain, KF7, as a potent lactic-acid-assimilating yeast. Heterothallic haploid strains KF7-5C and KF7-4B were obtained from the tetrads of the homothallic yeast strain KF7. The inefficient sporulation and poor spore viability of the haploid strains were improved by two strategies. The first strategy was as follows: (i) the KF7-5C was crossed with the laboratory strain SH6710; (ii) the progenies were backcrossed with KF7-5C three times; and (iii) the progenies were inbred three times to maintain a genetic background close to that of KF7. The NAM12 diploid between the cross of the resultant two strains, NAM11-9C and NAM11-13A, showed efficient sporulation and exhibited excellent growth in YPD medium (pH 3.5) at 35°C with 1.4-h generation time, indicating thermotolerance and acid tolerance. The second strategy was successive intrastrain crosses. The resultant two strains, KFG4-6B and KFG4-4B, showed excellent mating capacity. A spontaneous mutant of KFG4-6B, KFG4-6BD, showed a high growth rate with a generation time of 1.1 h in YPD medium (pH 3.0) at 35°C. The KFG4-6BD strain produced ascospores, which were crossed with NAM11-2C and its progeny to produce tetrads. These tetrads were crossed with KFG4-4B to produce NAM26-14A and NAM26-15A. The latter strain had a generation time of 1.6 h at 35°C in pH 2.5, thus exhibiting further thermotolerance and acid tolerance. A progeny from a cross of NAM26-14A and NAM26-15A yielded the strain NAM34-4C, which showed potent lactic acid assimilation and high transformation efficiency, better than those of a standard laboratory strain. Copyright © 2013 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

The impact of maternal HBsAg carrier status on pregnancy outcomes: a case-control study.

PubMed

Tse, Ka Yu; Ho, Lai Fong; Lao, Terence

2005-11-01

To examine the impact of maternal HBsAg carrier status on pregnancy outcomes. Two hundred and fifty-three carriers of hepatitis B surface antigen (HBsAg) with singleton pregnancy, were retrospectively compared with 253 controls matched for age and parity and year of delivery. On univariable analysis, HBsAg carriers had higher incidences of threatened preterm labour at <37 weeks (11.9% vs. 6.3%, P=0.030), preterm birth at <34 weeks (4.7% vs. 1.2%, P=0.033), gestational diabetes mellitus (19.0% vs. 11.1%, P=0.012) and antepartum haemorrhage (11.5% vs. 5.5%, P=0.026). Their infants had lower Apgar scores at the 1st (8.47+/-1.67 vs. 8.87+/-1.07, P=0.001) and 5th minute (9.56+/-1.29 vs. 9.80+/-0.54, P=0.007), and increased incidence of intraventricular haemorrhage (4.7% vs. 0.8%, P=0.007). On multivariable analysis, the association between HBsAg carrier state with antepartum haemorrhage, gestational diabetes mellitus and threatened preterm labour were confirmed. HBsAg carriers have increased risk of gestational diabetes mellitus, antepartum haemorrhage, and threatened preterm labour. This may be related to the chronic inflammatory state in these subjects. The role of chronic HBV infection in pregnancy complications has to be further elucidated.

Epidemiological characteristics of the carriers with coexistence of HBsAg and anti-HBs based on a community cohort study.

PubMed

Pu, Z; Li, D; Wang, A; Su, H; Shao, Z; Zhang, J; Ji, Z; Gao, J; Choi, B C K; Yan, Y

2016-04-01

The coexistence of HBsAg and anti-HBs is an atypical serological pattern in HBV infection. There is no epidemiological characteristics of this serological pattern in the community and there is controversy over the molecular mechanisms underlying this pattern. We investigated the epidemiological characteristics of the carriers with HBsAg and anti-HBs in a longitudinal community cohort study. The prevalence of this atypical serological pattern was 2.93% (122/4169) in HBsAg-positive populations. The prevalence progressively increased with age from 40 to 70 years old. The rate of HBeAg positive and detectable HBV DNA were both significantly higher in carriers with this pattern than in carriers who were HBsAg positive but anti-HBs negative (26/122 verse 598/4047, P = 0.046; 86/122 verse 275/529,P < 0.001). After 1 year of follow-up, 85.19% of the carriers still had coexistence HBsAg and anti-HBs, 14.81% of the carriers lost their anti-HBs. Viral sequencing showed that carriers with coexistence of HBsAg and anti-HBs had higher numbers of residue changes within the S gene than carriers who were HBsAg positive but anti-HBs negative (2.42 verse 1.33 changes per 100 residues, P < 0.05). Hence, the coexistence of HBsAg and anti-HBs is a unique serological pattern which may be associated with an increased risk of adverse clinical outcome and may be related to HBsAg immune variants which have genotypic heterogeneity. © 2015 John Wiley & Sons Ltd.

The anthracenedione compound bostrycin induces mitochondria-mediated apoptosis in the yeast Saccharomyces cerevisiae.

PubMed

Xu, Chunling; Wang, Jiafeng; Gao, Ye; Lin, Huangyu; Du, Lin; Yang, Shanshan; Long, Simei; She, Zhigang; Cai, Xiaoling; Zhou, Shining; Lu, Yongjun

2010-05-01

Bostrycin is an anthracenedione with phytotoxic and antibacterial activity that belongs to the large family of quinones. We have isolated bostrycin from the secondary metabolites of a mangrove endophytic fungus, no. 1403, collected from the South China Sea. Using the yeast Saccharomyces cerevisiae as a model, we show that bostrycin inhibits cell proliferation by blocking the cell cycle at G1 phase and ultimately leads to cell death in a time- and dose-dependent manner. Bostrycin-induced lethal cytotoxicity is accompanied with increased levels of intracellular reactive oxygen species and hallmarks of apoptosis such as chromatin condensation, DNA fragmentation and externalization of phosphatidylserine. We further show that bostrycin decreases mitochondrial membrane electric potential and causes mitochondrial destruction during the progression of cell death. Bostrycin-induced cell death was promoted in YCA1 null yeast strain but was partially rescued in AIF1 null mutant both in fermentative and respiratory media, strongly indicating that bostrycin induces apoptosis in yeast cells through a mitochondria-mediated but caspase-independent pathway.

Evaluation of HBsAg and anti-HBc assays in saliva and dried blood spot samples according HIV status.

PubMed

Flores, Geane Lopes; Cruz, Helena Medina; Potsch, Denise Vigo; May, Silvia Beatriz; Brandão-Mello, Carlos Eduardo; Pires, Marcia Maria Amendola; Pilotto, Jose Henrique; Lewis-Ximenez, Lia Laura; Lampe, Elisabeth; Villar, Livia Melo

2017-09-01

Influence of HIV status in HBV markers detection in saliva and dried blood spots (DBS) was not well established. This study aims to evaluate the performance of optimized commercial immunoassay for identifying HBsAg and anti-HBc in saliva and DBS according HIV status. A sum of 535 individuals grouped as HIV + , HBV + , HIV/HBV + and HIV/HBV- were recruited where 347 and 188 were included for HBsAg and anti-HBc evaluation, respectively. Serum, DBS collected in Whatman 903 paper and saliva obtained using salivette device were analyzed using EIA. Increased sample volume and ROC curve analysis for cut off determination were used for DBS and saliva testing. HBsAg detection in saliva and DBS exhibited sensitivities of 80.9% and 85.6% and specificities of 86.8% and 96.3%. Sensitivity of anti-HBc in saliva and DBS were 82.4% and 76.9% and specificities in saliva and DBS were 96.9% and 91.7%. Low sensitivities were observed for HBsAg (62%) and anti-HBc (47%) detection in saliva of HIV/HBV+ individuals. OD values were also lower for HBsAg detection in DBS and saliva of HIV/HBV+ individuals compared to their serum samples. Statistical significance was found for sensitivities in HBsAg detection between saliva and DBS demonstrating high sensitivity for DBS specimens. In conclusion, HIV status or antiretroviral treatment appears to interfere in the performance of HBsAg and anti-HBc detection in DBS and saliva samples using the adapted commercial EIA. Copyright © 2017 Elsevier B.V. All rights reserved.

Nationwide seroepidemiology of hepatitis B virus infection in South Korea in 2009 emphasizes the coexistence of HBsAg and anti-HBs.

PubMed

Lee, Byung Seok; Cho, Yong Kyun; Jeong, Sook-Hyang; Lee, Joon Hyeok; Lee, Don; Park, Neung Hwa; Ki, Moran

2013-08-01

Hepatitis B virus (HBV) infection is the major cause of chronic liver disease in Korea. This study investigated the seroprevalence of HBV infection with an emphasis on the coexistence of hepatitis B surface antigen (HBsAg) and antibody (anti-HBs). In all, 290,212 people undergoing health check-up examinations in 29 institutions during 2009 were recruited. The crude seroprevalences of HBsAg and anti-HBs was adjusted by age, sex, and geographic area using the 2009 estimated population of Korea. The adjusted seroprevalences of HBsAg and anti-HBs was 4.0% and 73.5%, respectively. Males showed higher HBsAg positivity and lower anti-HBs positivity than females (P<0.001). HBsAg positivity increased with age from 3.5% in people 20-29 years old to 4.8% in people 40-49 years old, followed by a decrease in people ≥ 50 years old. HBsAg positivity in Southern provinces (4.5%) including Jeju (5.9%), was significantly higher than that in Central provinces (3.6%; P<0.001). Interestingly, HBsAg and anti-HBs coexisted in 0.1% of the total subjects and in 2.9% of the HBsAg-positive group, showing distinct age distribution and higher alanine aminotransferase levels than those of the group positive for only HBsAg. In conclusion, the seroprevalence of HBsAg and anti-HBs in Korea varies significantly by age, sex and geographical location and coexisted in 2.9% of HBsAg-positive subjects. Continuous monitoring of seroepidemiology may facilitate the eventual eradication of HBV infection. Copyright © 2013 Wiley Periodicals, Inc.

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

Green tea polyphenols as potent enhancers of glucocorticoid-induced mouse mammary tumor virus gene expression.

PubMed

Abe, I; Umehara, K; Morita, R; Nemoto, K; Degawa, M; Noguchi, H

2001-02-16

The effect of natural and synthetic galloyl esters on glucocorticoid-induced gene expression was evaluated by using rat fibroblast 3Y1 cells stably transfected with a luciferase reporter gene under the transcriptional regulation of the mouse mammary tumor virus promoter. The glucocorticoid-induced gene transcription was strongly suppressed by synthetic alkyl esters; n-dodecyl gallate showed the most potent inhibition (66% inhibition at 10 microM), which was far more potent than that of crude tannic acid. n-Octyl and n-cetyl gallate also showed good inhibition, while gallic acid itself was not so active, suggesting that the presence of hydrophobic side chain is important for the suppressive effect. On the other hand, surprisingly, green tea gallocatechins, (-)-epigallocatechin-3-O-gallate and theasinensin A, potently enhanced the promoter activity (182 and 247% activity at 1 microM, respectively). The regulation of the level of the glucocorticoid-induced gene expression by the antioxidative gallates is of great interest from a therapeutic point of view.

Quantification of hepatitis B surface antigen with the novel DiaSorin LIAISON XL Murex HBsAg Quant: correlation with the ARCHITECT quantitative assays.

PubMed

Burdino, Elisa; Ruggiero, Tina; Proietti, Alex; Milia, Maria Grazia; Olivero, Antonella; Caviglia, Gian Paolo; Marietti, Milena; Rizzetto, Mario; Smedile, Antonina; Ghisetti, Valeria

2014-08-01

Recent technologic innovations allow for quantitative assessment of hepatitis B surface antigen (HBsAg) levels in serum; this has been used to monitor the course of chronic HBV hepatitis (CHB) and predict treatment response. LIAISON-XL Murex HBsAg Quant assay (DiaSorin, Saluggia, I) is the newest immunoassay CE approved to quantify HBsAg. To compare LIAISON-XL performances with ARCHITECT-QT HBsAg (Abbott Diagnostics, IL, USA), as reference test. Sequential serum samples (n=152) from 14 HBe-negative patients with CHB, the majority of them infected by HBV genotype D undergoing antiviral treatment, were retrospectively tested with both assays. The 2nd WHO Standard 00/588 for HBsAg was used as reference. LIAISON-XL and ARCHITECT-QT correlated by r=0.95, p<0.0001; by Bland-Altman analysis agreement of mean difference was 0.21 ± 0.15 log 10 IU/mL, 95% CI: -0.07 to 0.5). Performance of LIAISON-XL against the 2nd WHO Standard was r=0.998, p<0.0001 (95% CI: 0.993-0.999) with results nearer to the expected WHO values compared to ARCHITECT-QT. Median baseline HBsAg level was similar with the two methods before antiviral treatment, throughout fluctuations of HBsAg level in treatment non-responders and during the decrease of HBsAg titer in treatment responders. Correlation between HBsAg levels and HBV DNA was statistically significant for both the two immunoassays (LIAISON-XL: r=0.4988, 95% CI: 0.3452-0.6264, p<0.0001; ARCHITECT-QT: r=0.480, 95% CI: 0.3233-0.6111, p<0.0001). Correlation between HBsAg measurement with LIAISON-XL and ARCHITECT-QT was high. LIAISON-XL accurately quantified HBsAg in clinical samples at baseline or during antiviral therapy; it can be applied for HBsAg quantification in clinical practice and decision making in CHB. Copyright © 2014 Elsevier B.V. All rights reserved.

HBsAg quantification to predict natural history and treatment outcome in chronic hepatitis B patients.

PubMed

Martinot-Peignoux, Michelle; Asselah, Tarik; Marcellin, Patrick

2013-08-01

There is a growing interest in serum HBsAg quantification (qHbsAg). HBsAg titers are negatively correlated with liver fibrosis in HBeAg(+) patients. In HBeAg(-) HBsAg level <1000 IU/ml and HBV-DNA titer <2000 IU/ml accurately identify inactive carriers. During PEG-IFN treatment qHBsAg identifies patients with no benefit from therapy at week 12, allowing stopping or switched- "week 12 stopping rule". During nucleos(t)ide analogues the role of qHBsAg need to be clarified. In clinical practice qHBsAg is a simple and reproducible tool that may be used in association with HBV-DNA to classify patients during the natural history of HBV and to monitor therapy. Copyright © 2013 Elsevier Inc. All rights reserved.

HBsAg Loss with Peg-interferon Alfa-2a in Hepatitis B Patients with Partial Response to Nucleos(t)ide Analog: New Switch Study.

PubMed

Hu, Peng; Shang, Jia; Zhang, Wenhong; Gong, Guozhong; Li, Yongguo; Chen, Xinyue; Jiang, Jianning; Xie, Qing; Dou, Xiaoguang; Sun, Yongtao; Li, Yufang; Liu, Yingxia; Liu, Guozhen; Mao, Dewen; Chi, Xiaoling; Tang, Hong; Li, Xiaoou; Xie, Yao; Chen, Xiaoping; Jiang, Jiaji; Zhao, Ping; Hou, Jinlin; Gao, Zhiliang; Fan, Huimin; Ding, Jiguang; Zhang, Dazhi; Ren, Hong

2018-03-28

Background and Aims: Hepatitis B surface antigen (HBsAg) loss is seldom achieved with nucleos(t)ide analog (NA) therapy in chronic hepatitis B patients but may be enhanced by switching to finite pegylated-interferon (Peg-IFN) alfa-2a. We assessed HBsAg loss with 48- and 96-week Peg-IFN alfa-2a in chronic hepatitis B patients with partial response to a previous NA. Methods: Hepatitis B e antigen (HBeAg)-positive patients who achieved HBeAg loss and hepatitis B virus DNA <200 IU/mL with previous adefovir, lamivudine or entecavir treatment were randomized 1:1 to receive Peg-IFN alfa-2a for 48 ( n = 153) or 96 weeks ( n = 150). The primary endpoint of this study was HBsAg loss at end of treatment. The ClinicalTrials.gov identifier is NCT01464281. Results: At the end of 48 and 96 weeks' treatment, 14.4% (22/153) and 20.7% (31/150) of patients, respectively, who switched from NA to Peg-IFN alfa-2a cleared HBsAg. Rates were similar irrespective of prior NA or baseline HBeAg seroconversion. Among those who cleared HBsAg by the end of 48 and 96 weeks' treatment, 77.8% (14/18) and 71.4% (20/28), respectively, sustained HBsAg loss for a further 48 weeks. Baseline HBsAg <1500 IU/mL and week 24 HBsAg <200 IU/mL were associated with the highest rates of HBsAg loss at the end of both 48- and 96-week treatment (51.4% and 58.7%, respectively). Importantly, extending treatment from 48 to 96 weeks enabled 48.3% (14/29) more patients to achieve HBsAg loss. Conclusions: Patients on long-term NA who are unlikely to meet therapeutic goals can achieve high rates of HBsAg loss by switching to Peg-IFN alfa-2a. HBsAg loss rates may be improved for some patients by extending treatment from 48 to 96 weeks, although the differences in our study cohort were not statistically significant. Baseline and on-treatment HBsAg may predict HBsAg loss with Peg-IFN alfa-2a.

HBsAg plasma level kinetics: a new role for an old marker as a therapy response predictor in vertically infected children on combination therapy.

PubMed

Carey, I; Bruce, M; Horner, M; Zen, Y; D'Antiga, L; Bansal, S; Vergani, D; Mieli-Vergani, G

2015-04-01

We aimed to investigate the ability of HBsAg plasma level kinetics to predict therapy response by studying 23 children with infancy-acquired chronic hepatitis B (CHB) during combination sequential therapy with lead-in lamivudine (LAM) and add-on interferon-α (IFN-α) [5 responders (R = anti-HBs seroconversion) and 18 nonresponders (NR)] and to assess their relationship with pretreatment intrahepatic HBV-DNA and cccDNA and HBsAg and HBcAg liver expression. Plasma HBsAg levels were measured in samples before (treatment week 0 = TW0), during (TW9, TW28, TW52) and after (follow-up week = FUW24) therapy by Abbott ARCHITECT(®) assay [log10 IU/mL]. Baseline liver HBV-DNA and cccDNA were quantified by real-time TaqMan PCR [log10 copies/ng genomic DNA]. HBsAg and HBcAg liver expression was evaluated by immunostaining of formalin-fixed, paraffin-embedded specimens [number of positive cells/1000 hepatocytes]. All results are presented as medians. Plasma: at baseline, on-treatment and during follow-up, HBsAg levels were lower in R than NR (TW0: 4.36 vs 4.75;TW28: 2.44 vs 4.35;TW52: 0 vs 4.08 and FUW24: 0.17 vs 4.35, all P < 0.05). Liver: baseline HBV-DNA (3.82 vs 4.71, P = 0.16) and cccDNA (1.98 vs 2.26, P = 0.18) tended to be lower in R than NR, HBsAg expression was lower in R than NR (0.5 vs 4.7, P = 0.03), and HBcAg expression was similar between R and NR. There were positive correlations between plasma HBsAg levels and liver HBV-DNA (r = 0.44, P = 0.04), cccDNA (r = 0.41, P = 0.04) and HBsAg liver expression (r = 0.38, P = 0.05). Lower baseline HBsAg plasma levels, lower HBsAg expression in liver and on-treatment decline of plasma HBsAg levels heralds HBsAg clearance and response to treatment in tolerant children with CHB. © 2014 John Wiley & Sons Ltd.

Tim18, a component of the mitochondrial translocator, mediates yeast cell death induced by arsenic.

PubMed

Du, Li; Yu, Yong; Li, Zidong; Chen, Jingsi; Liu, Yan; Xia, Yongjing; Liu, Xiangjun

2007-08-01

Evidence is presented that Tim18, a mitochondria translocase, plays a role in the previously described apoptosis induced by arsenite in Saccharomyces cerevisiae. Tim18 deletion mutant exhibited resistance to arsenite. After arsenite treatment, both the wild type and Tim18-deficient cells showed reactive oxygen species (ROS) production. Arsenite induced the higher expression of tim18 in wild type yeast cells. We found that the tim18 deletion mutant also exhibited resistance to other apoptotic stresses such as acetic acid, H2O2, and hyperosmotic stress. These results suggest that Tim18 is important for yeast cell death induced by arsenic, and it may act downstream of ROS production.

Clinical, biochemical, virological and sonographic profile of incidentally detected asymptomatic HBsAg positive subjects, in Bangladesh.

PubMed

Ishaque, S M; Mahmuduzzaman, M; Rahman, M A; Uddoula, M S; Rahman, M Z; Khan, M R; Chowdhury, M S

2014-01-01

Chronic hepatitis B virus (HBV) is known to be the significant cause of Liver related morbidity and mortality, affecting 400 million people worldwide and a major public health problem in Bangladesh where carrier rates of HBV infection varies from 7.5 to 10%. In Bangladesh prevalence of asymptomatic HBV infection and incidentally detected HBsAg positive subjects were not well studied. The aim of this study is to evaluate the disease activity, replicative status of the virus and to find out the stages of chronic liver disease among incidentally detected asymptomatic HBsAg positive Bangladeshi subjects. Two hundred (200) incidentally detected healthy HBsAg positive subject were evaluated clinically, biochemically, serologically and ultrasonographically from January 2004 to June 2008. HBeAg was found positive in 17(8.5%), anti-HBe was positive in 174(87%), raised serum ALT (>45iu/L) in 45(22.5%), prothrombine time (PT) >3 sec of control in 33(16.5%). Ultrasonography showed coarse hepatic echotexture in 13(6.5%). Evidence of active viral replication and signs of chronic liver disease were observed among incidentally detected healthy HBsAg positive subjects. Such individuals should be followed up at regular interval to evaluate the replicative status of the virus and disease activity so that appropriate measures could be initiated in time.

Detection of a premature stop codon in the surface gene of hepatitis B virus from an HBsAg and antiHBc negative blood donor.

PubMed

Datta, Sibnarayan; Banerjee, Arup; Chandra, Partha K; Chakraborty, Subhasis; Basu, Subir Kumar; Chakravarty, Runu

2007-11-01

In blood donors, HBV infection is detected by the presence of serum hepatitis B surface antigen (HBsAg). However, some mutations in the surface gene region may result in altered or truncated HBsAg that can escape from immunoassay-based diagnosis. Such diagnostic escape mutants pose a potential risk for blood transfusion services. In the present study, we report a blood donor seronegative for HBsAg and antiHBc, but positive for antiHBs who was HBV DNA positive by PCR. Sequencing of the HBsAg gene revealed presence of a point mutation (T-A) at 207th nucleotide of the HBsAg ORF, which resulted in a premature stop codon at position 69. This results in a truncated HBsAg gene lacking the entire 'a' determinant region. However, follow-up of the donor after 2 years revealed clearance of HBV DNA from the serum. The case illustrates an unusual mutation, which causes HBsAg negativity. The finding emphasizes the importance of molecular assays in reducing the possibility of HBV transmission through blood transfusion. However, developing more sensitive serological assays, capable of detecting HBV mutants, is an alternative to expensive and complex amplification-based assays for developing countries.

Allicin disrupts the cell's electrochemical potential and induces apoptosis in yeast.

PubMed

Gruhlke, Martin C H; Portz, Daniela; Stitz, Michael; Anwar, Awais; Schneider, Thomas; Jacob, Claus; Schlaich, Nikolaus L; Slusarenko, Alan J

2010-12-15

The volatile substance allicin gives crushed garlic (Allium sativum) its characteristic odor and is a pro-oxidant that undergoes thiol-disulfide exchange reactions with -SH groups in proteins and glutathione. The antimicrobial activity of allicin is suspected to be due to the oxidative inactivation of essential thiol-containing enzymes. We investigated the hypothesis that at threshold inhibitory levels allicin can shunt yeast cells into apoptosis by altering their overall redox status. Yeast cells were treated either with chemically synthesized, pure allicin or with allicin in garlic juice. Allicin-dependent cell oxidation was demonstrated with a redox-sensitive GFP construct and the shift in cellular electrochemical potential (E(hc)) from less than -215 to -181mV was calculated using the Nernst equation after the glutathione/glutathione disulfide couple (2GSH/GSSG) in the cell was quantified. Caspase activation occurred after allicin treatment, and yeast expressing a human antiapoptotic Bcl-XL construct was rendered more resistant to allicin. Also, a yeast apoptosis-inducing factor deletion mutant was more resistant to allicin than wild-type cells. We conclude that allicin in garlic juice can activate apoptosis in yeast cells through its oxidizing properties and that this presents an alternative cell-killing mechanism to the previously proposed specific oxidative inactivation of essential enzymes. Copyright © 2010 Elsevier Inc. All rights reserved.

Acetate but not propionate induces oxidative stress in bakers' yeast Saccharomyces cerevisiae.

PubMed

Semchyshyn, Halyna M; Abrat, Oleksandra B; Miedzobrodzki, Jacek; Inoue, Yoshiharu; Lushchak, Volodymyr I

2011-01-01

The influence of acetic and propionic acids on baker's yeast was investigated in order to expand our understanding of the effect of weak organic acid food preservatives on eukaryotic cells. Both acids decreased yeast survival in a concentration-dependent manner, but with different efficiencies. The acids inhibited the fluorescein efflux from yeast cells. The inhibition constant of fluorescein extrusion from cells treated with acetate was significantly lower in parental strain than in either PDR12 (ABC-transporter Pdr12p) or WAR1 (transcriptional factor of Pdr12p) defective mutants. The constants of inhibition by propionate were virtually the same in all strains used. Yeast exposure to acetate increased the level of oxidized proteins and the activity of antioxidant enzymes, while propionate did not change these parameters. This suggests that various mechanisms underlie the yeast toxicity by acetic and propionic acids. Our studies with mutant cells clearly indicated the involvement of Yap1p transcriptional regulator and de novo protein synthesis in superoxide dismutase up-regulation by acetate. The up-regulation of catalase was Yap1p independent. Yeast pre-incubation with low concentrations of H₂O₂ caused cellular cross-protection against high concentrations of acetate. The results are discussed from the point of view that acetate induces a prooxidant effect in vivo, whereas propionate does not.

Hepatitis B virus infections in families in which the mothers are negative but the fathers are positive for HBsAg.

PubMed

Takegoshi, Kunio; Zhang, Wei

2006-10-01

We studied a total of 37 families, in which HBsAg was positive in either or both of father and mother, to assess intra-familial transmission of hepatitis B virus (HBV). The HBsAg positive rate for children with HBsAg-negative mothers was significantly lower than that with positive mothers (4 of 31, 12.9% versus 18 of 32, 56.3%, p<0.01) of course. However, there were three families in which the infection source for children was thought to be fathers, not mothers, i.e., of eight children in these three families with HBsAg +/- father/mother pairs, 4 (50%) were positive for both HBsAg and HBV DNA of genotypes identical to those of their fathers, and another child was positive for HBcAb despite being negative for HBsAg. Interestingly, moreover, all the mothers in these three families were HBcAb-positive even though HBsAg-negative, suggesting that not only father-to-child but also inter-spouse HBV transmission might have occurred. With these findings we would suggest that all the family members with HBsAg-positive fathers should receive HBV vaccine, let alone for those with HBsAg-positive mothers.

Wild yeast harbor a variety of distinct amyloid structures with strong prion-inducing capabilities

PubMed Central

Westergard, Laura; True, Heather L.

2014-01-01

Summary Variation in amyloid structures profoundly influences a wide array of pathological phenotypes in mammalian protein conformation disorders and dominantly inherited phenotypes in yeast. Here, we describe, for the first time, naturally occurring, self-propagating, structural variants of a prion protein isolated from wild strains of the yeast Saccharomyces cerevisiae. Variants of the [RNQ+] prion propagating in a variety of wild yeast differ biochemically, in their intracellular distributions, and in their ability to promote formation of the [PSI+] prion. [PSI+] is an epigenetic regulator of cellular phenotype and adaptability. Strikingly, we find that most natural [RNQ+] variants induced [PSI+] at high frequencies and the majority of [PSI+] variants elicited strong cellular phenotypes. We hypothesize that the presence of an efficient [RNQ+] template primes the cell for [PSI+] formation in order to induce [PSI+] in conditions where it would be advantageous. These studies utilize naturally occurring structural variants to expand our understanding of the consequences of diverse prion conformations on cellular phenotypes. PMID:24673812

Biosynthesized silver and gold nanoparticles are potent antimycotics against opportunistic pathogenic yeasts and dermatophytes.

PubMed

Rónavári, Andrea; Igaz, Nóra; Gopisetty, Mohana Krishna; Szerencsés, Bettina; Kovács, Dávid; Papp, Csaba; Vágvölgyi, Csaba; Boros, Imre Miklós; Kónya, Zoltán; Kiricsi, Mónika; Pfeiffer, Ilona

2018-01-01

Epidemiologic observations indicate that the number of systemic fungal infections has increased significantly during the past decades, however in human mycosis, mainly cutaneous infections predominate, generating major public health concerns and providing much of the impetus for current attempts to develop novel and efficient agents against cutaneous mycosis causing species. Innovative, environmentally benign and economic nanotechnology-based approaches have recently emerged utilizing principally biological sources to produce nano-sized structures with unique antimicrobial properties. In line with this, our aim was to generate silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs) by biological synthesis and to study the effect of the obtained nanoparticles on cutaneous mycosis causing fungi and on human keratinocytes. Cell-free extract of the red yeast Phaffia rhodozyma proved to be suitable for nanoparticle preparation and the generated AgNPs and AuNPs were characterized by transmission electron microscopy, dynamic light scattering and X-ray powder diffraction. Antifungal studies demonstrated that the biosynthesized silver particles were able to inhibit the growth of several opportunistic Candida or Cryptococcus species and were highly potent against filamentous Microsporum and Trichophyton dermatophytes. Among the tested species only Cryptococcus neoformans was susceptible to both AgNPs and AuNPs. Neither AgNPs nor AuNPs exerted toxicity on human keratinocytes. Our results emphasize the therapeutic potential of such biosynthesized nanoparticles, since their biocompatibility to skin cells and their outstanding antifungal performance can be exploited for topical treatment and prophylaxis of superficial cutaneous mycosis.

Tributyltin induces Yca1p-dependent cell death of yeast Saccharomyces cerevisiae.

PubMed

Chahomchuen, Thippayarat; Akiyama, Koichi; Sekito, Takayuki; Sugimoto, Naoko; Okabe, Masaaki; Nishimoto, Sogo; Sugahara, Takuya; Kakinuma, Yoshimi

2009-10-01

Tributyltin chloride (TBT), an environmental pollutant, is toxic to a variety of eukaryotic and prokaryotic organisms. Although it has been reported that TBT induces apoptotic cell death in mammalian, the action of TBT on eukaryotic microorganisms has not yet been fully investigated. In this study we examined the mechanism involved in cell death caused by TBT exposure in Saccharomyces cerevisiae. The median lethal concentration of TBT was 10 microM for the parent strain BY4741 and 3 microM for the pdr5Delta mutant defective in a major multidrug transporter, respectively. Fluorescence microscopic observations revealed nuclear condensation and chromatin fragmentation in cells treated with TBT indicating that cells underwent an apoptosis-like cell dearth. TBT-induced cell death was suppressed by deletion of the yca1 gene encoding a homologue of the mammalian caspase. In parallel, reactive oxygen species (ROS) were produced by TBT. These results suggest that TBT induces apoptosis-like cell death in yeast via an Yca1p-dependent pathway possibly downstream of the ROS production. This is the first report on TBT-induced apoptotic cell death in yeast.

Discovery of a diaminoquinoxaline benzenesulfonamide antagonist of HIV-1 Nef function using a yeast-based phenotypic screen

PubMed Central

2013-01-01

Background HIV-1 Nef is a viral accessory protein critical for AIDS progression. Nef lacks intrinsic catalytic activity and binds multiple host cell signaling proteins, including Hck and other Src-family tyrosine kinases. Nef binding induces constitutive Hck activation that may contribute to HIV pathogenesis by promoting viral infectivity, replication and downregulation of cell-surface MHC-I molecules. In this study, we developed a yeast-based phenotypic screen to identify small molecules that inhibit the Nef-Hck complex. Results Nef-Hck interaction was faithfully reconstituted in yeast cells, resulting in kinase activation and growth arrest. Yeast cells expressing the Nef-Hck complex were used to screen a library of small heterocyclic compounds for their ability to rescue growth inhibition. The screen identified a dihydrobenzo-1,4-dioxin-substituted analog of 2-quinoxalinyl-3-aminobenzene-sulfonamide (DQBS) as a potent inhibitor of Nef-dependent HIV-1 replication and MHC-I downregulation in T-cells. Docking studies predicted direct binding of DQBS to Nef which was confirmed in differential scanning fluorimetry assays with recombinant purified Nef protein. DQBS also potently inhibited the replication of HIV-1 NL4-3 chimeras expressing Nef alleles representative of all M-group HIV-1 clades. Conclusions Our findings demonstrate the utility of a yeast-based growth reversion assay for the identification of small molecule Nef antagonists. Inhibitors of Nef function discovered with this assay, such as DQBS, may complement the activity of current antiretroviral therapies by enabling immune recognition of HIV-infected cells through the rescue of cell surface MHC-I. PMID:24229420

Novel structural features in Candida albicans hyphal glucan provide a basis for differential innate immune recognition of hyphae versus yeast.

PubMed

Lowman, Douglas W; Greene, Rachel R; Bearden, Daniel W; Kruppa, Michael D; Pottier, Max; Monteiro, Mario A; Soldatov, Dmitriy V; Ensley, Harry E; Cheng, Shih-Chin; Netea, Mihai G; Williams, David L

2014-02-07

The innate immune system differentially recognizes Candida albicans yeast and hyphae. It is not clear how the innate immune system effectively discriminates between yeast and hyphal forms of C. albicans. Glucans are major components of the fungal cell wall and key fungal pathogen-associated molecular patterns. C. albicans yeast glucan has been characterized; however, little is known about glucan structure in C. albicans hyphae. Using an extraction procedure that minimizes degradation of the native structure, we extracted glucans from C. albicans hyphal cell walls. (1)H NMR data analysis revealed that, when compared with reference (1→3,1→6) β-linked glucans and C. albicans yeast glucan, hyphal glucan has a unique cyclical or "closed chain" structure that is not found in yeast glucan. GC/MS analyses showed a high abundance of 3- and 6-linked glucose units when compared with yeast β-glucan. In addition to the expected (1→3), (1→6), and 3,6 linkages, we also identified a 2,3 linkage that has not been reported previously in C. albicans. Hyphal glucan induced robust immune responses in human peripheral blood mononuclear cells and macrophages via a Dectin-1-dependent mechanism. In contrast, C. albicans yeast glucan was a much less potent stimulus. We also demonstrated the capacity of C. albicans hyphal glucan, but not yeast glucan, to induce IL-1β processing and secretion. This finding provides important evidence for understanding the immune discrimination between colonization and invasion at the mucosal level. When taken together, these data provide a structural basis for differential innate immune recognition of C. albicans yeast versus hyphae.

Novel Structural Features in Candida albicans Hyphal Glucan Provide a Basis for Differential Innate Immune Recognition of Hyphae Versus Yeast*

PubMed Central

Lowman, Douglas W.; Greene, Rachel R.; Bearden, Daniel W.; Kruppa, Michael D.; Pottier, Max; Monteiro, Mario A.; Soldatov, Dmitriy V.; Ensley, Harry E.; Cheng, Shih-Chin; Netea, Mihai G.; Williams, David L.

2014-01-01

The innate immune system differentially recognizes Candida albicans yeast and hyphae. It is not clear how the innate immune system effectively discriminates between yeast and hyphal forms of C. albicans. Glucans are major components of the fungal cell wall and key fungal pathogen-associated molecular patterns. C. albicans yeast glucan has been characterized; however, little is known about glucan structure in C. albicans hyphae. Using an extraction procedure that minimizes degradation of the native structure, we extracted glucans from C. albicans hyphal cell walls. 1H NMR data analysis revealed that, when compared with reference (1→3,1→6) β-linked glucans and C. albicans yeast glucan, hyphal glucan has a unique cyclical or “closed chain” structure that is not found in yeast glucan. GC/MS analyses showed a high abundance of 3- and 6-linked glucose units when compared with yeast β-glucan. In addition to the expected (1→3), (1→6), and 3,6 linkages, we also identified a 2,3 linkage that has not been reported previously in C. albicans. Hyphal glucan induced robust immune responses in human peripheral blood mononuclear cells and macrophages via a Dectin-1-dependent mechanism. In contrast, C. albicans yeast glucan was a much less potent stimulus. We also demonstrated the capacity of C. albicans hyphal glucan, but not yeast glucan, to induce IL-1β processing and secretion. This finding provides important evidence for understanding the immune discrimination between colonization and invasion at the mucosal level. When taken together, these data provide a structural basis for differential innate immune recognition of C. albicans yeast versus hyphae. PMID:24344127

A pathway of targeted autophagy is induced by DNA damage in budding yeast

PubMed Central

Eapen, Vinay V.; Waterman, David P.; Bernard, Amélie; Schiffmann, Nathan; Sayas, Enrich; Kamber, Roarke; Lemos, Brenda; Memisoglu, Gonen; Ang, Jessie; Mazella, Allison; Chuartzman, Silvia G.; Loewith, Robbie J.; Schuldiner, Maya; Denic, Vladimir; Klionsky, Daniel J.; Haber, James E.

2017-01-01

Autophagy plays a central role in the DNA damage response (DDR) by controlling the levels of various DNA repair and checkpoint proteins; however, how the DDR communicates with the autophagy pathway remains unknown. Using budding yeast, we demonstrate that global genotoxic damage or even a single unrepaired double-strand break (DSB) initiates a previously undescribed and selective pathway of autophagy that we term genotoxin-induced targeted autophagy (GTA). GTA requires the action primarily of Mec1/ATR and Rad53/CHEK2 checkpoint kinases, in part via transcriptional up-regulation of central autophagy proteins. GTA is distinct from starvation-induced autophagy. GTA requires Atg11, a central component of the selective autophagy machinery, but is different from previously described autophagy pathways. By screening a collection of ∼6,000 yeast mutants, we identified genes that control GTA but do not significantly affect rapamycin-induced autophagy. Overall, our findings establish a pathway of autophagy specific to the DNA damage response. PMID:28154131

A pathway of targeted autophagy is induced by DNA damage in budding yeast.

PubMed

Eapen, Vinay V; Waterman, David P; Bernard, Amélie; Schiffmann, Nathan; Sayas, Enrich; Kamber, Roarke; Lemos, Brenda; Memisoglu, Gonen; Ang, Jessie; Mazella, Allison; Chuartzman, Silvia G; Loewith, Robbie J; Schuldiner, Maya; Denic, Vladimir; Klionsky, Daniel J; Haber, James E

2017-02-14

Autophagy plays a central role in the DNA damage response (DDR) by controlling the levels of various DNA repair and checkpoint proteins; however, how the DDR communicates with the autophagy pathway remains unknown. Using budding yeast, we demonstrate that global genotoxic damage or even a single unrepaired double-strand break (DSB) initiates a previously undescribed and selective pathway of autophagy that we term genotoxin-induced targeted autophagy (GTA). GTA requires the action primarily of Mec1/ATR and Rad53/CHEK2 checkpoint kinases, in part via transcriptional up-regulation of central autophagy proteins. GTA is distinct from starvation-induced autophagy. GTA requires Atg11, a central component of the selective autophagy machinery, but is different from previously described autophagy pathways. By screening a collection of ∼6,000 yeast mutants, we identified genes that control GTA but do not significantly affect rapamycin-induced autophagy. Overall, our findings establish a pathway of autophagy specific to the DNA damage response.

Membranous glomerulonephritis in a child asymptomatic for hepatitis B virus. Concomitant seropositivity for HBsAG and anti-HBs.

PubMed

Hirsch, H Z; Ainsworth, S K; DeBeukelaer, M; Brissie, R M; Hennigar, G R

1981-04-01

The presence of hepatitis B surface antigen (HBsAg) in association with immunoglobulins and complement components within the glomerular basement membranes of adults having chronic active hepatitis has been well documented. In addition, investigators in Poland have demonstrated HBsAg immune complexes in glomeruli of children who did not have clinical evidence of hepatitis. More recently, a single case of childhood membranous glomerulonephritis in an asymptomatic carrier of hepatitis B virus was cited by observers in Canada. Reported here is the deposition of HBsAg immune complexes in the glomerular basement membranes of a 13-year-old black boy who had membranous glomerulopathy but not clinical evidence of hepatitis. This may be the first reported case in the United States of HbsAg-associated membranous glomerulonephritis in a child asymptomatic for hepatitis B virus, and only the second such case in North America. However, unlike previous studies of childhood glomerulopathy in association with hepatitis B virus, this patient is seropositive for both HBsAg and anti-HBs (antibody for hepatitis B surface antigen). Similar "rare" serologic findings were found for the patient's eldest male sib.

HBsAg carrier status and the association between gestational diabetes with increased serum ferritin concentration in Chinese women.

PubMed

Lao, Terence T; Tse, Ka-Yu; Chan, Louis Y; Tam, Kar-Fai; Ho, Lai-Fong

2003-11-01

To determine whether the high prevalence of hepatitis B surface antigen (HBsAg) carriage in our population can explain the previous observation of an association between increased maternal serum ferritin concentration and gestational diabetes in Hong Kong Chinese women. A retrospective study was performed on 767 nonanemic women with singleton pregnancy who had iron status assessed at 28-30 weeks. The result of the routine antenatal HBsAg screening was retrieved from patient records. The HBsAg-positive and -negative groups were compared for maternal characteristics, prevalence of gestational diabetes in the third trimester, prevalence of high serum ferritin and iron concentrations, and transferrin saturation, which is defined as a value in the highest quartile established by the measurements obtained from the HBsAg-negative group. The incidences of oral glucose tolerance test and gestational diabetes were significantly increased in the HBsAg-positive group. The HBsAg-positive women with gestational diabetes had significantly increased prevalence of high serum ferritin compared with the HBsAg-negative women, irrespective of the latter's gestational diabetes status. Multiple logistic regression analysis confirmed the independent association between HBsAg carrier status with gestational diabetes (relative risk 3.51, 95% CI 1.83-6.73) but excluded high ferritin as an independent factor. Our results indicate that maternal HBsAg carriage could explain in part the association between increased serum ferritin concentration with gestational diabetes in Hong Kong Chinese women, and that HBsAg carrier status is an independent risk factor for gestational diabetes.

Luteolin-7-O-Glucoside Present in Lettuce Extracts Inhibits Hepatitis B Surface Antigen Production and Viral Replication by Human Hepatoma Cells in Vitro

PubMed Central

Cui, Xiao-Xian; Yang, Xiao; Wang, Hui-Jing; Rong, Xing-Yu; Jing, Sha; Xie, You-Hua; Huang, Dan-Feng; Zhao, Chao

2017-01-01

Hepatitis B virus (HBV) infection is endemic in Asia and chronic hepatitis B (CHB) is a major public health issue worldwide. Current treatment strategies for CHB are not satisfactory as they induce a low rate of hepatitis B surface antigen (HBsAg) loss. Extracts were prepared from lettuce hydroponically cultivated in solutions containing glycine or nitrate as nitrogen sources. The lettuce extracts exerted potent anti-HBV effects in HepG2 cell lines in vitro, including significant HBsAg inhibition, HBV replication and transcription inhibition, without exerting cytotoxic effects. When used in combination interferon-alpha 2b (IFNα-2b) or lamivudine (3TC), the lettuce extracts synergistically inhibited HBsAg expression and HBV replication. By using differential metabolomics analysis, Luteolin-7-O-glucoside was identified and confirmed as a functional component of the lettuce extracts and exhibited similar anti-HBV activity as the lettuce extracts in vitro. The inhibition rate on HBsAg was up to 77.4%. Moreover, both the lettuce extracts and luteolin-7-O-glucoside functioned as organic antioxidants and, significantly attenuated HBV-induced intracellular reactive oxygen species (ROS) accumulation. Luteolin-7-O-glucoside also normalized ROS-induced mitochondrial membrane potential damage, which suggests luteolin-7-O-glucoside inhibits HBsAg and HBV replication via a mechanism involving the mitochondria. Our findings suggest luteolin-7-O-glucoside may have potential value for clinical application in CHB and may enhance HBsAg and HBV clearance when used as a combination therapy. PMID:29270164

Lack of chemically induced mutation in repair-deficient mutants of yeast.

PubMed

Prakash, L

1974-12-01

Two genes, rad6 and rad9, that confer radiation sensitivity in the yeast Saccharomyces cerevisiae also greatly reduce the frequency of chemically-induced reversions of a tester mutant cyc1-131, which is a chain initiation mutant in the structural gene determining iso-1-cytochrome c. Mutations induced by ethyl methanesulfonate (EMS), diethyl sulfate (DES), methyl methanesulfonate (MMS), dimethyl sulfate (DMS), nitroquinoline oxide (NQO), nitrosoguanidine (NTG), nitrogen mustard (HN2), beta-propiolactone, and tritiated uridine, as well as mutations induced by ultraviolet light (UV) and ionizing radiation were greatly diminished in strains homozygous for either the rad6 or rad9 gene. Nitrous acid and nitrosoimidazolidone (NIL), on the other hand, were highly mutagenic in these repair-deficient mutants, and at low doses, these mutagens acted with about the same efficiency as in the normal RAD strain. At high doses of either nitrous acid or NIL, however, reversion frequencies were significantly reduced in the two rad mutants compared to normal strains. Although both rad mutants are immutable to about the same extent, the rad9 strains tend to be less sensitive to the lethal effect of chemical mutagens than rad6 strains. It is concluded that yeast requires a functional repair system for mutation induction by chemical agents.

Conditionally controlling nuclear trafficking in yeast by chemical-induced protein dimerization

PubMed Central

Xu, Tao; Johnson, Cole A; Gestwicki, Jason E; Kumar, Anuj

2016-01-01

We present here a protocol to conditionally control the nuclear trafficking of target proteins in yeast. In this system, rapamycin is used to heterodimerize two chimeric proteins. one chimera consists of a FK506-binding protein (FKBp12) fused to a cellular ‘address’ (nuclear localization signal or nuclear export sequence). the second chimera consists of a target protein fused to a fluorescent protein and the FKBp12-rapamycin-binding (FrB) domain from FKBp-12-rapamycin associated protein 1 (Frap1, also known as mtor). rapamycin induces dimerization of the FKBp12- and FrB-containing chimeras; these interactions selectively place the target protein under control of the cell address, thereby directing the protein into or out of the nucleus. By chemical-induced dimerization, protein mislocalization is reversible and enables the identification of conditional loss-of-function and gain-of-function phenotypes, in contrast to other systems that require permanent modification of the targeted protein. Yeast strains for this analysis can be constructed in 1 week, and the technique allows protein mislocalization within 15 min after drug treatment. PMID:21030958

Conditionally controlling nuclear trafficking in yeast by chemical-induced protein dimerization.

PubMed

Xu, Tao; Johnson, Cole A; Gestwicki, Jason E; Kumar, Anuj

2010-11-01

We present here a protocol to conditionally control the nuclear trafficking of target proteins in yeast. In this system, rapamycin is used to heterodimerize two chimeric proteins. One chimera consists of a FK506-binding protein (FKBP12) fused to a cellular 'address' (nuclear localization signal or nuclear export sequence). The second chimera consists of a target protein fused to a fluorescent protein and the FKBP12-rapamycin-binding (FRB) domain from FKBP-12-rapamycin associated protein 1 (FRAP1, also known as mTor). Rapamycin induces dimerization of the FKBP12- and FRB-containing chimeras; these interactions selectively place the target protein under control of the cell address, thereby directing the protein into or out of the nucleus. By chemical-induced dimerization, protein mislocalization is reversible and enables the identification of conditional loss-of-function and gain-of-function phenotypes, in contrast to other systems that require permanent modification of the targeted protein. Yeast strains for this analysis can be constructed in 1 week, and the technique allows protein mislocalization within 15 min after drug treatment.

Yeast-incorporated gallium attenuates glucocorticoid-induced bone loss in rats by inhibition of bone resorption.

PubMed

Ren, Zhaozhou; Yang, Liqing; Xue, Feng; Meng, Qingjie; Wang, Kejia; Wu, Xian; Ji, Chao; Jiang, Teng; Liu, Da; Zhou, Long; Zhang, Jing; Fu, Qin

2013-06-01

Glucocorticoids (GC) are potent anti-inflammatory agents and widely used for the treatment of many immune-mediated and inflammatory diseases, whereas GC-induced osteoporosis (GIOP) is the most common cause of secondary osteoporosis and significantly increases the patients' morbidity and mortality. GIOP is characterized as diminished osteogenesis and accelerated bone resorption. Yeast-incorporated gallium (YG) as an organic compound not only reduces elements-associated toxicity, but also maintains its therapeutic effect on improving bone loss or promoting fracture healing in ovariectomized female rats. The aim of this study was to examine whether YG could prevent GC-induced bone loss. Five-month-old male Sprague-Dawley rats were randomly divided into three groups (n = 6): two groups were administered dexamethasone (0.1 mg/kg/day) or vehicle (PBS) subcutaneously for 5 weeks; one other group was received dexamethasone subcutaneously and YG (120 μg/kg/day) orally. Trabecular bone microarchitectural parameters, bone mineral density (BMD), bone strength, body weight, and serum biochemical markers of bone resorption and formation were examined. Compared to the GC alone group, treatment with YG not only prevented microarchitectural deterioration of trabecular bone volume relative to tissue volume, trabecular number, and trabecular separation, but also significantly improved BMD, mechanical strength, and body weight in GC-treated rats. Moreover, YG decreased tartrate-resistant acid phosphatase 5b level but failed to change alkaline phosphatase level in GC-treated rats. This is the first study to show that YG prominently attenuates bone loss and microarchitectural deterioration and inhibits the increased bone resorption in GIOP. It implies that YG might be an alternative therapy for prevention of GC-induced bone loss in humans.

Clinical performance of the novel DiaSorin LIAISON(®) XL murex: HBsAg Quant, HCV-Ab, HIV-Ab/Ag assays.

PubMed

Krawczyk, Adalbert; Hintze, Christian; Ackermann, Jessica; Goitowski, Birgit; Trippler, Martin; Grüner, Nico; Neumann-Fraune, Maria; Verheyen, Jens; Fiedler, Melanie

2014-01-01

The fully automated and closed LIAISON(®)XL platform was developed for reliable detection of infection markers like hepatitis B virus (HBV) surface antigen (HBsAg), hepatitis C virus (HCV) antibodies (Ab) or human immunodeficiency virus (HIV)-Ag/Ab. To date, less is known about the diagnostic performance of this system in direct comparison to the common Abbott ARCHITECT(®) platform. We compared the diagnostic performance and usability of the DiaSorin LIAISON(®)XL with the commonly used Abbott ARCHITECT(®) system. The qualitative performance of the above mentioned assays was compared in about 500 sera. Quantitative tests were performed for HBsAg-positive samples from patients under therapy (n=289) and in vitro expressed mutants (n=37). For HCV-Ab, a total number of 155 selected samples from patients chronically infected with different HCV genotypes were tested. The concordance between both systems was 99.4% for HBsAg, 98.81% for HCV-Ab, and 99.6% for HIV-Ab/Ag. The quantitative LIAISON(®)XL murex HBsAg assay detected all mutants in comparable amounts to the HBsAg wild type and yielded highly reliable HBsAg kinetics in patients treated with antiviral drugs. Dilution experiments using the 2nd International Standard for HBsAg (WHO) showed a high accuracy of this test. HCV-Ab from patients infected with genotypes 1-3 were equally detected in both systems. Interestingly, S/CO levels of HCV-Ab from patients infected with genotype 3 seem to be relatively low using both systems. The LIAISON(®)XL platform proved to be an excellent system for diagnostics of HBV, HCV, and HIV with equal performance compared to the ARCHITECT(®) system. Copyright © 2013 Elsevier B.V. All rights reserved.

[Intragenic mitotic recombination induced by ultraviolet and gamma rays in radiosensitive mutants of Saccharomyces cerevisiae yeasts].

PubMed

Zakharov, I A; Kasinova, G V; Koval'tsova, S V

1983-01-01

The effect of UV- and gamma-irradiation on the survival and intragenic mitotic recombination (gene conversion) of 5 radiosensitive mutants was studied in comparison with the wild type. The level of spontaneous conversion was similar for RAD, rad2 and rad15, mutations xrs2 and xrs4 increasing and rad54 significantly decreasing it. The frequency of conversion induced by UV-light was greater in rad2, rad15 and xrs2 mutants and lower in xrs4, as compared to RAD. Gamma-irradiation caused induction of gene conversion with an equal frequency in RAD, rad2, rad15. Xrs2 and xrs4 mutations slightly decreased gamma-induced conversion. In rad54 mutant, UV-and gamma-induced conversion was practically absent. In the wild type yeast, a diploid strain is more resistant than a haploid, whereas in rad54 a diploid strain has the same or an increased sensitivity, as compared to a haploid strain (the "inverse ploidy effect"). This effect and also the block of induced mitotic recombination caused by rad54 indicate the presence in the yeast Saccharomyces cerevisiae of repair pathways of UV- and gamma-induced damages acting in diploid cells and realised by recombination. The data obtained as a result of many years' investigation of genetic effects in radiosensitive mutants of yeast are summarised and considered.

The phytopathogenic virulent effector protein RipI induces apoptosis in budding yeast Saccharomyces cerevisiae.

PubMed

Deng, Meng-Ying; Sun, Yun-Hao; Li, Pai; Fu, Bei; Shen, Dong; Lu, Yong-Jun

2016-10-01

Virulent protein toxins secreted by the bacterial pathogens can cause cytotoxicity by various molecular mechanisms to combat host cell defense. On the other hand, these proteins can also be used as probes to investigate the defense pathway of host innate immunity. Ralstonia solanacearum, one of the most virulent bacterial phytopathogens, translocates more than 70 effector proteins via type III secretion system during infection. Here, we characterized the cytotoxicity of effector RipI in budding yeast Saccharomyce scerevisiae, an alternative host model. We found that over-expression of RipI resulted in severe growth defect and arginine (R) 117 within the predicted integrase motif was required for inhibition of yeast growth. The phenotype of death manifested the hallmarks of apoptosis. Our data also revealed that RipI-induced apoptosis was independent of Yca1 and mitochondria-mediated apoptotic pathways because Δyca1 and Δaif1 were both sensitive to RipI as compared with the wild type. We further demonstrated that RipI was localized in the yeast nucleus and the N-terminal 1-174aa was required for the localization. High-throughput RNA sequencing analysis showed that upon RipI over-expression, 101 unigenes of yeast ribosome presented lower expression level, and 42 GO classes related to the nucleus or recombination were enriched with differential expression levels. Taken together, our data showed that a nuclear-targeting effector RipI triggers yeast apoptosis, potentially dependent on its integrase function. Our results also provided an alternative strategy to dissect the signaling pathway of cytotoxicity induced by the protein toxins. Copyright © 2016 Elsevier Ltd. All rights reserved.

Clinical Significance of Quantitative HBsAg Titres and its Correlation With HBV DNA Levels in the Natural History of Hepatitis B Virus Infection.

PubMed

Karra, Vijay K; Chowdhury, Soumya J; Ruttala, Rajesh; Polipalli, Sunil K; Kar, Premashis

2016-09-01

Quantification of serum hepatitis B antigen (HBsAg) is an important test that marks active infection with hepatitis B and helps in the prediction of the clinical outcome and management of hepatitis B virus (HBV) infection. Correlation with HBV DNA quantitative levels may help in developing strategies for antiviral treatment. This study is aimed to evaluate HBsAg titres in various phase of HBV infection in HBsAg positive patients, and its correlation with HBV DNA viral load levels. 976 HBV related patients were analysed in this retrospective cross-sectional study. Patients were categorised on the basis of the phase of HBV infection: immune tolerant phase (IT, n  = 123), immune clearance phase (IC, n  = 192), low-replicative phase (LR, n  = 476), and HBeAg-negative hepatitis (ENH, n  = 185). HBsAg titres were quantified and correlated with HBV-DNA levels and clinical parameters. Median HBsAg titres were different between each phases of HBV infection ( P  < 0.001): (4.62 log10 IU/ml), IC (3.88 log10 IU/ml), LR (2.76 log10 IU/ml) and ENH (2.94 log10 IU/ml). HBsAg and HBV DNA levels showed significant correlation in the whole group ( r  = 0.694, P  < 0.001), and this was also observed in different phases of HBV infection. Strong correlation in IT phase ( r  = 0.603, P  < 0.001) and IC phase ( r  = 0.523, P  < 0.001), moderate in LR phase ( r  = 0.362, P  < 0.001) and weak in ENH ( r  = 0.110, P  = 0.04). No correlation was observed between serum HBsAg levels and biochemical parameters. The study demonstrated significant difference in the median baseline values of serum HBsAg titres in different phases of HBV infection and provides additional information in understanding the natural history of HBV-infection.

Purification of HBsAg produced by the human hepatoma cell line PLC/PRE/5 by affinity chromatography using monoclonal antibodies and application for ELISA diagnostic.

PubMed

Merten, O W; Reiter, S; Scheirer, W; Katinger, H

1983-01-01

The human cell line PLC/PRF/5 (5) was used for the production of hepatitis B surface antigen subtype ad (HBsAg ad) and purified by affinity chromatography (AC) with monoclonal antibodies (mAb). mAb to HBsAg from mouse ascites have been purified by Protein A - AC prior coupling to AH-Sepharose 4B (Pharmacia). The combined procedure of ammonium-sulphate-precipitation of HBsAg from culture supernatants and immunosorbent-AC leads to approx. 700-fold purification. ELISA results using the mAb and the HBsAg for diagnostics of human serum, positive for anti-HBsAg-antibodies correlate with the RIA (AUSAB, Abbott).

Not your ordinary yeast: non-Saccharomyces yeasts in wine production uncovered.

PubMed

Jolly, Neil P; Varela, Cristian; Pretorius, Isak S

2014-03-01

Saccharomyces cerevisiae and grape juice are 'natural companions' and make a happy wine marriage. However, this relationship can be enriched by allowing 'wild' non-Saccharomyces yeast to participate in a sequential manner in the early phases of grape must fermentation. However, such a triangular relationship is complex and can only be taken to 'the next level' if there are no spoilage yeast present and if the 'wine yeast' - S. cerevisiae - is able to exert its dominance in time to successfully complete the alcoholic fermentation. Winemakers apply various 'matchmaking' strategies (e.g. cellar hygiene, pH, SO2 , temperature and nutrient management) to keep 'spoilers' (e.g. Dekkera bruxellensis) at bay, and allow 'compatible' wild yeast (e.g. Torulaspora delbrueckii, Pichia kluyveri, Lachancea thermotolerans and Candida/Metschnikowia pulcherrima) to harmonize with potent S. cerevisiae wine yeast and bring the best out in wine. Mismatching can lead to a 'two is company, three is a crowd' scenario. More than 40 of the 1500 known yeast species have been isolated from grape must. In this article, we review the specific flavour-active characteristics of those non-Saccharomyces species that might play a positive role in both spontaneous and inoculated wine ferments. We seek to present 'single-species' and 'multi-species' ferments in a new light and a new context, and we raise important questions about the direction of mixed-fermentation research to address market trends regarding so-called 'natural' wines. This review also highlights that, despite the fact that most frontier research and technological developments are often focussed primarily on S. cerevisiae, non-Saccharomyces research can benefit from the techniques and knowledge developed by research on the former. © 2013 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Identified OAS3 gene variants associated with coexistence of HBsAg and anti-HBs in chronic HBV infection.

PubMed

Wang, S; Wang, J; Fan, M-J; Li, T-Y; Pan, H; Wang, X; Liu, H-K; Lin, Q-F; Zhang, J-G; Guan, L-P; Zhernakova, D V; O'Brien, S J; Feng, Z-R; Chang, L; Dai, E-H; Lu, J-H; Xi, H-L; Zeng, Z; Yu, Y-Y; Wang, B-B

2018-03-27

The underlying mechanism of coexistence of hepatitis B surface antigen (HBsAg) and hepatitis B surface antigen antibody (anti-HBs) is still controversial. To identify the host genetic factors related to this unusual clinical phenomenon, a two-stage study was conducted in the Chinese Han population. In the first stage, we performed a case-control (1:1) age- and gender-matched study of 101 cases with concurrent HBsAg and anti-HBs and 102 controls with negative HBsAg and positive anti-HBs using whole exome sequencing. In the second validation stage, we directly sequence the 16 exons on the OAS3 gene in two dependent cohorts of 48 cases and 200 controls. Although, in the first stage, a genome-wide association study of 58,563 polymorphism variants in 101 cases and 102 controls found no significant loci (P-value ≤ .05/58563), and neither locus achieved a conservative genome-wide significance threshold (P-value ≤ 5e-08), gene-based burden analysis showed that OAS3 gene rare variants were associated with the coexistence of HBsAg and anti-HBs. (P-value = 4.127e-06 ≤ 0.05/6994). A total of 16 rare variants were screened out from 21 cases and 3 controls. In the second validation stage, one case with a stop-gained rare variant was identified. Fisher's exact test of all 149 cases and 302 controls showed that the rare coding sequence mutations were more frequent in cases vs controls (P-value = 7.299e-09, OR = 17.27, 95% CI [5.01-58.72]). Protein-coding rare variations on the OAS3 gene are associated with the coexistence of HBsAg and anti-HBs in patients with chronic HBV infection in Chinese Han population. © 2018 John Wiley & Sons Ltd.

Tributyltin induces cell cycle arrest at G1 phase in the yeast Saccharomyces cerevisiae.

PubMed

Sekito, Takayuki; Sugimoto, Naoko; Ishimoto, Masaya; Kawano-Kawada, Miyuki; Akiyama, Koichi; Nishimoto, Sogo; Sugahara, Takuya; Kakinuma, Yoshimi

2014-04-01

Tributyltin (TBT) has long been recognized as a major environmental pollutant that can cause significant damage to the cellular functions as well as disruption of endocrine homeostasis. TBT induces apoptosis accompanied by production of reactive oxygen species (ROS) in mammalian and yeast cells. We observed that the budding yeast cells exposed to this compound at low concentrations exhibited cell growth arrest, but not cell death. Flow cytometric analysis of yeast cells without synchronization and morphological assessment of cells synchronized at M phase by nocodazole treatment indicated that TBT-exposed Saccharomyces cerevisiae cells were arrested at G1 phase of the cell cycle. This arrest was recovered by the addition of N-acetylcysteine, suggesting the involvement of ROS production by TBT. This is the first study to evaluate the action of TBT on cell cycle events.

Protective effect of Brewer's yeast on methimazole-induced-adrenal atrophy (a stereological study).

PubMed

Dehghani, Farzaneh; Zabolizadeh, Jamal; Noorafshan, Ali; Panjehshahin, Mohammad Reza; Karbalay-Doust, Saied

2010-04-20

Induction of hypothyroidism by thioamide drugs will cause adrenal gland atrophy and decrease in its hormones. To prevent side effect on the adrenal gland, brewer's yeast, a natural product rich in vitamins and minerals was used. Serological techniques were applied to measure the volume of adrenal gland. For this purpose, 48 Sprague-Dawley rats were randomly divided into one control and three experimental groups. In group 1, methimazole was administered at the dose of 30 mg/kg/day days, in group 2, 120 mg/kg/day of, brewer's yeast, in group 3, 30 mg/kg/day of methimazole plus 120 mg/kg/day of brewer yeast, and for the control group, an equal volume of saline (0.5 ml/rat/day) was orally given. After 30 days, all the animals were anesthetized and their adrenal glands were removed, fixed, embedded and stained. The volume of different zones of the adrenal glands was estimated by Cavalieri principle and point counting methods. statistical analysis was performed using Mann-Withney test and p < 0.05 was considered as statistically significant. The results indicated that methimazole decreased the volume of fasciculata zone in the cortex of the adrenal gland and also decreased the blood cortisol level. Brewer's yeast reduced the methimazole side effects on this zone. In conclusion, it seems that the use of brewer's yeast could prevent methimazole-induced atrophy of the adrenal gland.

New mutations affecting induced mutagenesis in yeast.

PubMed

Lawrence, C W; Krauss, B R; Christensen, R B

1985-01-01

Previously isolated mutations in baker's yeast, Saccharomyces cerevisiae, that impair induced mutagenesis were all identified with the aid of tests that either exclusively or predominantly detect base-pair substitutions. To avoid this bias, we have screened 11 366 potentially mutant clones for UV-induced reversion of the frameshift allele, his4-38, and have identified 10 mutants that give much reduced yields of revertants. Complementation and recombination tests show that 6 of these carry mutations at the previously known REV1, REV1 and REV3 loci, while the remaining 4 define 3 new genes, REV4 (2 mutations), REV5 and REV6. The rev4 mutations are readily suppressed in many genetic backgrounds and, like the rev5 mutation, impart only a limited deficiency for induced mutagenesis: it is likely, therefore that the REV4+ and REV5+ gene functions are only remotely concerned with this process. The rev6 mutants have a more general deficiency, however, as well as marked sensitivity to UV and an increased spontaneous mutation rate, properties that suggest the REV6 gene is directly involved in mutation induction. The REV5 gene is located about 1 cM proximal to CYC1 on chromosome X.

Heavy ion induced DNA-DSB in yeast and mammalian cells

NASA Technical Reports Server (NTRS)

Loebrich, M.; Ikpeme, S.; Kiefer, J.

1994-01-01

Molecular changes at the DNA are assumed to be the main cause for radiation effects in a number of organisms. During the course of the last decades techniques have been developed for measuring DNA double-strand breaks (dsb), generally assumed to be the most critical DNA lesions. The outcome of all those different approaches portrays a collection of data useful for a theoretical description of radiation action mechanisms. However, in the case of heavy ion induced DNA dsb the picture is not quite clear yet and further projects and strategies have to be developed. The biological systems studied in our group are yeast and mammalian cells. While in the case of yeast cells technical and methodical reasons highlight these organisms mammalian cells reach greater importance when dsb repair studies are performed. In both types of organisms the technique of pulsed-field gel electrophoresis (PFGE) is applied, although with different modifications and evaluation procedures mainly due to the different genome sizes.

Zinc starvation induces autophagy in yeast

PubMed Central

Kawamata, Tomoko; Horie, Tetsuro; Matsunami, Miou; Sasaki, Michiko; Ohsumi, Yoshinori

2017-01-01

Zinc is an essential nutrient for all forms of life. Within cells, most zinc is bound to protein. Because zinc serves as a catalytic or structural cofactor for many proteins, cells must maintain zinc homeostasis under severely zinc-deficient conditions. In yeast, the transcription factor Zap1 controls the expression of genes required for uptake and mobilization of zinc, but to date the fate of existing zinc-binding proteins under zinc starvation remains poorly understood. Autophagy is an evolutionarily conserved cellular degradation/recycling process in which cytoplasmic proteins and organelles are sequestered for degradation in the vacuole/lysosome. In this study, we investigated how autophagy functions under zinc starvation. Zinc depletion induced non-selective autophagy, which is important for zinc-limited growth. Induction of autophagy by zinc starvation was not directly related to transcriptional activation of Zap1. Instead, TORC1 inactivation directed zinc starvation-induced autophagy. Abundant zinc proteins, such as Adh1, Fba1, and ribosomal protein Rpl37, were degraded in an autophagy-dependent manner. But the targets of autophagy were not restricted to zinc-binding proteins. When cellular zinc is severely depleted, this non-selective autophagy plays a role in releasing zinc from the degraded proteins and recycling zinc for other essential purposes. PMID:28264932

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.

A synthetic chalcone as a potent inducer of glutathione biosynthesis.

PubMed

Kachadourian, Remy; Day, Brian J; Pugazhenti, Subbiah; Franklin, Christopher C; Genoux-Bastide, Estelle; Mahaffey, Gregory; Gauthier, Charlotte; Di Pietro, Attilio; Boumendjel, Ahcène

2012-02-09

Chalcones continue to attract considerable interest due to their anti-inflammatory and antiangiogenic properties. We recently reported the ability of 2',5'-dihydroxychalcone (2',5'-DHC) to induce both breast cancer resistance protein-mediated export of glutathione (GSH) and c-Jun N-terminal kinase-mediated increased intracellular GSH levels. Herein, we report a structure-activity relationship study of a series of 30 synthetic chalcone derivatives with hydroxyl, methoxyl, and halogen (F and Cl) substituents and their ability to increase intracellular GSH levels. This effect was drastically improved with one or two electrowithdrawing groups on phenyl ring B and up to three methoxyl and/or hydroxyl groups on phenyl ring A. The optimal structure, 2-chloro-4',6'-dimethoxy-2'-hydroxychalcone, induced both a potent NF-E2-related factor 2-mediated transcriptional response and an increased formation of glutamate cysteine ligase holoenzyme, as shown using a human breast cancer cell line stably expressing a luciferase reporter gene driven by antioxidant response elements.

Prediction of conformational changes by single mutation in the hepatitis B virus surface antigen (HBsAg) identified in HBsAg-negative blood donors.

PubMed

Ie, Susan I; Thedja, Meta D; Roni, Martono; Muljono, David H

2010-11-18

Selection of hepatitis B virus (HBV) by host immunity has been suggested to give rise to variants with amino acid substitutions at or around the 'a' determinant of the surface antigen (HBsAg), the main target of antibody neutralization and diagnostic assays. However, there have never been successful attempts to provide evidence for this hypothesis, partly because the 3 D structure of HBsAg molecules has not been determined. Tertiary structure prediction of HBsAg solely from its primary amino acid sequence may reveal the molecular energetic of the mutated proteins. We carried out this preliminary study to analyze the predicted HBsAg conformation changes of HBV variants isolated from Indonesian blood donors undetectable by HBsAg assays and its significance, compared to other previously-reported variants that were associated with diagnostic failure. Three HBV variants (T123A, M133L and T143M) and a wild type sequence were analyzed together with frequently emerged variants T123N, M133I, M133T, M133V, and T143L. Based on the Jameson-Wolf algorithm for calculating antigenic index, the first two amino acid substitutions resulted in slight changes in the antigenicity of the 'a' determinant, while all four of the comparative variants showed relatively more significant changes. In the pattern T143M, changes in antigenic index were more significant, both in its coverage and magnitude, even when compared to variant T143L. These data were also partially supported by the tertiary structure prediction, in which the pattern T143M showed larger shift in the HBsAg second loop structure compared to the others. Single amino acid substitutions within or near the 'a' determinant of HBsAg may alter antigenicity properties of variant HBsAg, which can be shown by both its antigenic index and predicted 3 D conformation. Findings in this study emphasize the significance of variant T143M, the prevalent isolate with highest degree of antigenicity changes found in Indonesian blood donors. This

Alpha-ketoglutarate enhances freeze-thaw tolerance and prevents carbohydrate-induced cell death of the yeast Saccharomyces cerevisiae.

PubMed

Bayliak, Maria M; Hrynkiv, Olha V; Knyhynytska, Roksolana V; Lushchak, Volodymyr I

2018-01-01

Stress resistance and fermentative capability are important quality characteristics of baker's yeast. In the present study, we examined protective effects of exogenous alpha-ketoglutarate (AKG), an intermediate of the tricarboxylic acid cycle and amino acid metabolism, against freeze-thaw and carbohydrate-induced stresses in the yeast Saccharomyces cerevisiae. Growth on AKG-supplemented medium prevented a loss of viability and improved fermentative capacity of yeast cells after freeze-thaw treatment. The cells grown in the presence of AKG had higher levels of amino acids (e.g., proline), higher metabolic activity and total antioxidant capacity, and higher activities of catalase, NADP-dependent glutamate dehydrogenase and glutamine synthase compared to control ones. Both synthesis of amino acids and enhancement of antioxidant system capacity could be involved in AKG-improved freeze-thaw tolerance in S. cerevisiae. Cell viability dramatically decreased under incubation of stationary-phase yeast cells in 2% glucose or fructose solutions (in the absence of the other nutrients) as compared with incubation in distilled water or in 10 mM AKG solution. The decrease in cell viability was accompanied by acidification of the medium, and decrease in cellular respiration, aconitase activity, and levels of total protein and free amino acids. The supplementation with 10 mM AKG effectively prevented carbohydrate-induced yeast death. Protective mechanisms of AKG could be associated with the intensification of respiration and prevention of decreasing protein level as well as with direct antioxidant AKG action.

Detection of HBV genome in the plasma and peripheral blood mononuclear cells of Iranian HBsAg negative patients with HIV infection: occult HBV infection.

PubMed

Tajik, Zahra; Bokharaei-Salim, Farah; Ghorbani, Saied; Keyvani, Hossein; Esghaei, Maryam; Monavari, Seyed Hamidreza; Ataei-Pirkooh, Angila; Garshasbi, Saba; Donyavi, Tahereh; Fakhim, Atousa

2018-06-01

The presence of hepatitis B virus (HBV) DNA in the absence of traceable hepatitis B surface antigen (HBsAg) in the plasma specimen of patients is defined as occult HBV infection (OBI). This study aimed to detect HBV-DNA in the plasma and peripheral blood mononuclear cells (PBMCs) of Iranian HBsAg negative patients with human immunodeficiency virus (HIV) infection. This cross-sectional study was conducted on 172 patients with HIV infection from September 2015 to August 2017. The patients were tested for serological parameters (HBsAg, HBcAb, HBeAg and HBeAb) against HBV infection. Moreover, they were tested for HBV viral load (using COBAS TaqMan 48 Kit, Roche, USA) in plasma and the presence of the HBV genome in PBMC specimens using real-time PCR. The mean age of the patients was 35.4 ± 13.4 years. Of the 172 studied patients, 109 (63.4%) were male. In this study, 151 (87.8%) patients were negative for HBsAg, 111 (64.5%) patients were negative for all HBV infection serological markers, 9 (5.2%) patients were only positive for HBsAg and 29 (16.9%) patients were only positive for HBcAb. Moreover, five (3.3%) patients with HBsAg negative had OBI (in the plasma sample of four patients and PBMC specimens of all five patients, HBV-DNA was detected). The present study revealed that 3.3% of the patients with HIV infection had occult HBV infection. Presumably, designing prospective studies to identify this infection in patients with HIV infection is informative and valuable.

Zinc starvation induces autophagy in yeast.

PubMed

Kawamata, Tomoko; Horie, Tetsuro; Matsunami, Miou; Sasaki, Michiko; Ohsumi, Yoshinori

2017-05-19

Zinc is an essential nutrient for all forms of life. Within cells, most zinc is bound to protein. Because zinc serves as a catalytic or structural cofactor for many proteins, cells must maintain zinc homeostasis under severely zinc-deficient conditions. In yeast, the transcription factor Zap1 controls the expression of genes required for uptake and mobilization of zinc, but to date the fate of existing zinc-binding proteins under zinc starvation remains poorly understood. Autophagy is an evolutionarily conserved cellular degradation/recycling process in which cytoplasmic proteins and organelles are sequestered for degradation in the vacuole/lysosome. In this study, we investigated how autophagy functions under zinc starvation. Zinc depletion induced non-selective autophagy, which is important for zinc-limited growth. Induction of autophagy by zinc starvation was not directly related to transcriptional activation of Zap1. Instead, TORC1 inactivation directed zinc starvation-induced autophagy. Abundant zinc proteins, such as Adh1, Fba1, and ribosomal protein Rpl37, were degraded in an autophagy-dependent manner. But the targets of autophagy were not restricted to zinc-binding proteins. When cellular zinc is severely depleted, this non-selective autophagy plays a role in releasing zinc from the degraded proteins and recycling zinc for other essential purposes. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Effects of the type III secreted pseudomonal toxin ExoS in the yeast Saccharomyces cerevisiae.

PubMed

Stirling, Fiona R; Evans, Tom J

2006-08-01

Pseudomonas aeruginosa secretes a number of toxins by a type III system, and these are important in virulence. One of them, ExoS, is a bifunctional toxin, with a GTPase-activating protein domain, as well as ADP ribosyltransferase (ADPRT) activity. These two domains have numerous potential cellular targets, but the overall mechanism of ExoS action remains unclear. The effects of ExoS in a simple eukaryotic system, the yeast Saccharomyces cerevisiae, using a tetracycline-regulated expression system were studied. This system allowed controlled expression of ExoS in yeast, which was not possible using a galactose-induced system. ExoS was found to be an extremely potent inhibitor of yeast growth, and to be largely dependent on the activity of its ADPRT domain. ExoS produced a dramatic alteration in actin distribution, with the appearance of large aggregates of cortical actin, and thickened disorganized cables, entirely dependent on the ADPRT domain. This phenotype is suggestive of actin stabilization, which was verified by showing that the cortical aggregates of actin induced by ExoS were resistant to treatment with latrunculin A, an agent that prevents actin polymerization. ExoS increased the numbers of mating projections produced following growth arrest with mating pheromone, and prevented subsequent DNA replication, an effect that is again dependent on the ADPRT domain. Following pheromone removal, ExoS produced altered development of the mating projections, which became elongated with a swollen bud-like tip. These results suggest alternative pathways for ExoS action in eukaryotic cells that may result from activation of small GTPases, and this yeast expression system is well suited to explore these pathways.

Accelerating Yeast Prion Biology using Droplet Microfluidics

NASA Astrophysics Data System (ADS)

Ung, Lloyd; Rotem, Assaf; Jarosz, Daniel; Datta, Manoshi; Lindquist, Susan; Weitz, David

2012-02-01

Prions are infectious proteins in a misfolded form, that can induce normal proteins to take the misfolded state. Yeast prions are relevant, as a model of human prion diseases, and interesting from an evolutionary standpoint. Prions may also be a form of epigenetic inheritance, which allow yeast to adapt to stressful conditions at rates exceeding those of random mutations and propagate that adaptation to their offspring. Encapsulation of yeast in droplet microfluidic devices enables high-throughput measurements with single cell resolution, which would not be feasible using bulk methods. Millions of populations of yeast can be screened to obtain reliable measurements of prion induction and loss rates. The population dynamics of clonal yeast, when a fraction of the cells are prion expressing, can be elucidated. Furthermore, the mechanism by which certain strains of bacteria induce yeast to express prions in the wild can be deduced. Integrating the disparate fields of prion biology and droplet microfluidics reveals a more complete picture of how prions may be more than just diseases and play a functional role in yeast.

Sero-prevalence of HBsAg in naive HIV-infected patients in a rural locality of Cameroon.

PubMed

Molu, Jean-Patrick; Essome, Marie Chantal Ngonde; Monamele, Chavely Gwladys; Njouom, Richard

2018-01-16

This study was performed in order to fill the gap of knowledge regarding sero-epidemiology of hepatitis B virus (HBV) amongst Human Immunodeficiency virus (HIV)-infected patients and to assess the risk factors associated with HBV co-infection in a rural locality of Cameroon. A retrospective and cross-sectional study was carried out from January 2008 to April 2014 within the Mfou District Hospital. Naive HIV-infected patients were enrolled in the study and tested for hepatitis B surface antigen (HBsAg). Preliminary pre-therapeutic data essential for follow-up was collected from the participants. Overall, the sample size was constituted of 712 HIV-infected patients. The prevalence of HBsAg was 8.99%. A significant difference was observed in the proportion of HBsAg positive subjects with respect to the year of inclusion; higher proportions were observed between 2011 and 2014 (P-value = 0.007). Majority of HBV co-infected participants had severe immuno-suppression with CD4 counts lower than 100 cells/µL as compared to HIV mono-infected population but the difference was not statistically significant. Our results confirm the high prevalence for HBV infection among HIV-infected patients in the Mfou District Hospital. These findings will enable stake holders to be better armed in the elimination of viral hepatitis as a public health problem.

Isolation of the alkane inducible cytochrome P450 (P450alk) gene from the yeast Candida tropicalis

EPA Science Inventory

The gene for the alkane-inducible cytochrome P450, P450alk, has been isolated from the yeast Candida tropicalis by immunoscreening a λgt11 library. Isolation of the gene has been identified on the basis of its inducibility and partial DNA sequence. Transcripts of this gene were i...

Nonmutagenic carcinogens induce intrachromosomal recombination in dividing yeast cells.

PubMed

Schiestl, R H

1993-12-01

A large number of animal and human carcinogens without apparent genotoxic activity exist (nonmutagenic carcinogens) that are difficult or impossible to detect with the currently used short-term tests. Because of the association of carcinogenesis with genome rearrangement, a system selecting for intrachromosomal recombination (DEL recombination) that results in genome rearrangement has been constructed in the yeast Saccharomyces cerevisiae. Because DEL recombination is under different genetic control than interchromosomal recombination and meiotic recombination, it is probably due to a different mechanism. It has been found that DEL recombination is readily inducible by 10 mutagenic carcinogens and 17 nonmutagenic carcinogens that are not detectable (false negatives) with the Ames assay. In addition, three out of four mutagens that do not cause cancer (false positives in the Ames assay) do not induce DEL recombination. DEL recombination is inducible by UV only in dividing cells but not in cells synchronized in the G1 or G2 phase of the cell cycle. Interchromosomal recombination, on the other hand, is inducible in G1 but not in G2. The nonmutagenic carcinogens induce DEL recombination only in actively growing cells, which may give some indication as to their mechanism. Further characterization of the mechanism involved in induction of DEL recombination may contribute to the understanding of the biological activity of nonmutagenic carcinogens.

K2 killer toxin-induced physiological changes in the yeast Saccharomyces cerevisiae.

PubMed

Orentaite, Irma; Poranen, Minna M; Oksanen, Hanna M; Daugelavicius, Rimantas; Bamford, Dennis H

2016-03-01

Saccharomyces cerevisiae cells produce killer toxins, such as K1, K2 and K28, that can modulate the growth of other yeasts giving advantage for the killer strains. Here we focused on the physiological changes induced by K2 toxin on a non-toxin-producing yeast strain as well as K1, K2 and K28 killer strains. Potentiometric measurements were adjusted to observe that K2 toxin immediately acts on the sensitive cells leading to membrane permeability. This correlated with reduced respiration activity, lowered intracellular ATP content and decrease in cell viability. However, we did not detect any significant ATP leakage from the cells treated by killer toxin K2. Strains producing heterologous toxins K1 and K28 were less sensitive to K2 than the non-toxin producing one suggesting partial cross-protection between the different killer systems. This phenomenon may be connected to the observed differences in respiratory activities of the killer strains and the non-toxin-producing strain at low pH. This might also have practical consequences in wine industry; both as beneficial ones in controlling contaminating yeasts and non-beneficial ones causing sluggish fermentation. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Berberine potentizes apoptosis induced by X-rays irradiation probably through modulation of gap junctions.

PubMed

Liu, Bing; Wang, Qin; Yuan, Dong-dong; Hong, Xiao-ting; Tao, Liang

2011-04-01

Clinical combination of some traditional Chinese medical herbs, including berberine, with irradiation is demonstrated to improve efficacy of tumor radiotherapy, yet the mechanisms for such effect remain largely unknown. The present study investigated the effect of berberine on apoptosis induced by X-rays irradiation and the relation between this effect and gap junction intercellular communication (GJIC). The role of gap junctions in the modulation of X-rays irradiation-induced apoptosis was explored by manipulation of connexin (Cx) expression, and gap junction function, using oleamide, a GJIC inhibitor, and berberine. In transfected HeLa cells, Cx32 expression increased apoptosis induced by X-rays irradiation, while inhibition of gap junction by oleamide reduced the irradiation responses, indicating the dependence of X-rays irradiation-induced apoptosis on GJIC. Berberine, at the concentrations without cytotoxicity, enhanced apoptosis induced by irradiation only in the presence of functional gap junctions. These results suggest that berberine potentizes cell apoptosis induced by X-rays irradiation, probably through enhancement of gap junction activity.

[Comparative sensitivity of different solid-phase immunoassays for detecting HBsAg and anti-HCV in blood].

PubMed

León, P; López, J A; Domingo, C J; Echevarría, J M

1992-10-01

The sensitivities of seven methods of enzyme-immunoassay for HBsAg detection, as well as of twelve immunoassays for detecting antibody to HCV, were compared, in an attempt to evaluate the need for an official technical validation of such methods prior to its commercialization in Spain. Significant differences were seen for the sensitivities of these methods, which may influence the proficiency of blood bank screening for HBsAg and anti-HCV for preventing cases of post-transfusional viral hepatitis. As a conclusion, it is recommended to establish legal regulations for pre-marketing validation of such assays in Spain, as well as to take the results obtained in these evaluation studies as the basis for selecting those tests which may be more convenient for screening purposes at the blood banks.

A Synthetic Chalcone as a Potent Inducer of Glutathione Biosynthesis

PubMed Central

Kachadourian, Remy; Day, Brian J.; Pugazhenti, Subbiah; Franklin, Christopher C.; Genoux-Bastide, Estelle; Mahaffey, Gregory; Gauthier, Charlotte; Di Pietro, Attilio; Boumendjel, Ahcène

2014-01-01

Chalcones continue to attract considerable interest due to their anti-inflammatory and antiangiogenic properties. We recently reported the ability of 2′,5′-dihydroxychalcone (2′,5′-DHC) to induce both breast cancer resistance protein-mediated export of glutathione (GSH) and c-Jun N-terminal kinase-mediated increased intracellular GSH levels. Herein, we report a structure–activity relationship study of a series of 30 synthetic chalcone derivatives with hydroxyl, methoxyl, and halogen (F and Cl) substituents and their ability to increase intracellular GSH levels. This effect was drastically improved with one or two electrowithdrawing groups on phenyl ring B and up to three methoxyl and/or hydroxyl groups on phenyl ring A. The optimal structure, 2-chloro-4′,6′-dimethoxy-2′-hydroxychalcone, induced both a potent NF-E2-related factor 2-mediated transcriptional response and an increased formation of glutamate cysteine ligase holoenzyme, as shown using a human breast cancer cell line stably expressing a luciferase reporter gene driven by antioxidant response elements. PMID:22239485

CKA2 functions in H2O2-induced apoptosis and high-temperature stress tolerance by regulating NO accumulation in yeast.

PubMed

Liu, Wen-Cheng; Yuan, Hong-Mei; Li, Yun-Hui; Lu, Ying-Tang

2015-09-01

Nitric oxide (NO) plays key roles in yeast responses to various environmental factors, such as H2O2 and high temperature. However, the gene encoding NO synthase (NOS) in yeast has not yet been identified, and the mechanism underlying the regulation of NOS-like activity is poorly understood. Here, we report on the involvement of CKA2 in H2O2-induced yeast apoptosis and yeast high-temperature stress tolerance. Our results showed that although Δcka2 mutant had reduced NO accumulation with decreased apoptosis after H2O2 exposure, treatment with a NO donor, sodium nitroprusside, resulted in similar survival rate of Δcka2 mutant compared to that of wild-type yeast when subjected to H2O2 stress. This finding occurred because H2O2-enhanced NOS-like activity in wild-type yeast was significantly repressed in Δcka2. Our additional experiments indicated that both high-temperature-enhanced NO accumulation and NOS-like activity were also suppressed in Δcka2, leading to the hypersensitivity of the mutant to high temperature in terms of changes in survival rate. Thus, our results showed that CKA2 functioned in H2O2-induced apoptosis and high-temperature stress tolerance by regulating NOS-like-dependent NO accumulation in yeast. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Yeast-derived Particulate β-Glucan Treatment Subverts the Suppression of Myeloid-derived Suppressor Cells by Inducing PMN-MDSC Apoptosis and M-MDSC Differentiation to APC in Cancer

PubMed Central

Albeituni, Sabrin H.; Ding, Chuanlin; Liu, Min; Hu, Xiaoling; Luo, Fengling; Kloecker, Goetz; Bousamra, Micahel; Zhang, Huang-ge; Yan, Jun

2016-01-01

Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of immature myeloid cells that promote tumor progression. Herein, we demonstrated that activation of a C-type lectin receptor, dectin-1, in MDSC differentially modulates the function of different MDSC subsets. Yeast-derived whole β-glucan particles (WGP), a ligand to engage and activate dectin-1, oral treatment in vivo significantly decreased tumor weight and splenomegaly in tumor-bearing mice with reduced accumulation of PMN-MDSC but not M-MDSC, and decreased PMN-MDSC suppression in vitro through the induction of respiratory burst and apoptosis. On a different axis, WGP-treated M-MDSC differentiated into F4/80+CD11c+ cells in vitro that served as potent antigen-presenting cells (APC) to induce Ag-specific CD4+ and CD8+ T cell responses in a dectin-1 dependent manner. In addition, ERK1/2 phosphorylation was required for the acquisition of APC properties in M-MDSC. Moreover, WGP-treated M-MDSC differentiated into CD11c+ cells in vivo with high MHC class II expression and induced decreased tumor burden when inoculated subcutaneously with LLC cells. This effect was dependent of the dectin-1 receptor. Strikingly, patients with non-small cell lung cancer (NSCLC) that had received WGP treatment for 10–14 days prior to any other treatment had a decreased frequency of CD14−HLA-DR−CD11b+CD33+ MDSC in the peripheral blood. Overall, these data indicate that WGP may be a potent immune modulator of MDSC suppressive function and differentiation in cancer. PMID:26810222

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

The Phospholipid:Diacylglycerol Acyltransferase Lro1 Is Responsible for Hepatitis C Virus Core-Induced Lipid Droplet Formation in a Yeast Model System

PubMed Central

Wang, Chao-Wen; Cheng, Yun-Hsin; Irokawa, Hayato; Hwang, Gi-Wook; Naganuma, Akira; Kuge, Shusuke

2016-01-01

Chronic infection with the hepatitis C virus frequently induces steatosis, which is a significant risk factor for liver pathogenesis. Steatosis is characterized by the accumulation of lipid droplets in hepatocytes. The structural protein core of the virus induces lipid droplet formation and localizes on the surface of the lipid droplets. However, the precise molecular mechanisms for the core-induced formation of lipid droplets remain elusive. Recently, we showed that the expression of the core protein in yeast as a model system could induce lipid droplet formation. In this study, we probed the cellular factors responsible for the formation of core-induced lipid-droplets in yeast cells. We demonstrated that one of the enzymes responsible for triglyceride synthesis, a phospholipid:diacylglycerol acyltransferase (Lro1), is required for the core-induced lipid droplet formation. While core proteins inhibit Lro1 degradation and alter Lro1 localization, the characteristic localization of Lro1 adjacent to the lipid droplets appeared to be responsible for the core-induced lipid droplet formation. RNA virus genomes have evolved using high mutation rates to maintain their ability to replicate. Our observations suggest a functional relationship between the core protein with hepatocytes and yeast cells. The possible interactions between core proteins and the endoplasmic reticulum membrane affect the mobilization of specific proteins. PMID:27459103

Historic and current hepatitis B viral DNA and quantitative HBsAg level are not associated with cirrhosis in non-Asian women with chronic hepatitis B.

PubMed

Harkisoen, S; Arends, J E; van den Hoek, J A R; Whelan, J; van Erpecum, K J; Boland, G J; Hoepelman, A I M

2014-12-01

Some studies done in Asian patients have shown that serum levels of hepatitis B virus (HBV) DNA predict the development of cirrhosis. However, it is unclear whether this also applies for non-Asian patients. This study investigated historic and current HBV DNA and quantitative hepatitis B surface antigen (HBsAg) levels as predictors of cirrhosis in non-Asian women with chronic HBV. A retrospective cohort study of non-Asian women with chronic HBV was performed. Among other variables, HBV DNA and quantitative HBsAg levels were measured in stored historic serum samples obtained during pregnancy (period 1990-2004) and current serum samples (period 2011-2012) to determine any association with liver cirrhosis by liver stiffness measurement (LSM). One hundred and nineteen asymptomatic, treatment-naïve non-Asian women were included; the median number of years between the historic sample and the current sample was 17 (interquartile range (IQR) 13-20). The median historic log HBV DNA and quantitative log HBsAg levels were 2.5 (IQR 1.9-3.4) IU/ml and 4.2 (IQR 3.6-4.5) IU/ml, respectively. LSM diagnosed 14 patients (12%) with F3-F4 fibrosis, i.e. stiffness >8.1kPa. No association of cirrhosis was found with historic HBV DNA (relative risk (RR) 0.34, 95% confidence interval (CI) 0.05-2.44) or with the quantitative HBsAg level (HBsAg level >1000 IU/ml, RR 0.35, 95% CI 0.11-1.11). Multivariable analysis identified alcohol consumption (odds ratio (OR) 6.4, 95% CI 1.3-30.1), aspartate aminotransferase >0.5 times the upper limit of normal (OR 15.4, 95% CI 1.9-122.6), and prothrombin time (OR 12.0, 95% CI 1.2-120.4), but not HBV DNA or quantitative HBsAg level, to be independent predictors of the presence of cirrhosis. Neither historic nor current HBV DNA or the quantitative HBsAg level is associated with the development of HBV-related cirrhosis in non-Asian women. Copyright © 2014 The Authors. Published by Elsevier Ltd.. All rights reserved.

Oral immunization with hepatitis B surface antigen expressed in transgenic plants

PubMed Central

Kong, Qingxian; Richter, Liz; Yang, Yu Fang; Arntzen, Charles J.; Mason, Hugh S.; Thanavala, Yasmin

2001-01-01

Oral immunogenicity of recombinant hepatitis B surface antigen (HBsAg) derived from yeast (purified product) or in transgenic potatoes (uncooked unprocessed sample) was compared. An oral adjuvant, cholera toxin, was used to increase immune responses. Transgenic plant material containing HBsAg was the superior means of both inducing a primary immune response and priming the mice to respond to a subsequent parenteral injection of HBsAg. Electron microscopy of transgenic plant samples revealed evidence that the HBsAg accumulated intracellularly; we conclude that natural bioencapsulation of the antigen may provide protection from degradation in the digestive tract until plant cell degradation occurs near an immune effector site in the gut. The correlate of protection from hepatitis B virus infection is serum antibody titers induced by vaccination; the protective level in humans is 10 milliunits/ml or greater. Mice fed HBsAg-transgenic potatoes produced HBsAg-specific serum antibodies that exceeded the protective level and, on parenteral boosting, generated a strong long-lasting secondary antibody response. We have also shown the effectiveness of oral delivery by using a parenteral prime-oral boost immunization schedule. The demonstrated success of oral immunization for hepatitis B virus with an “edible vaccine” provides a strategy for contributing a means to achieve global immunization for hepatitis B prevention and eradication. PMID:11553782

The Yeast Environmental Stress Response Regulates Mutagenesis Induced by Proteotoxic Stress

PubMed Central

Shor, Erika; Fox, Catherine A.; Broach, James R.

2013-01-01

Conditions of chronic stress are associated with genetic instability in many organisms, but the roles of stress responses in mutagenesis have so far been elucidated only in bacteria. Here, we present data demonstrating that the environmental stress response (ESR) in yeast functions in mutagenesis induced by proteotoxic stress. We show that the drug canavanine causes proteotoxic stress, activates the ESR, and induces mutagenesis at several loci in an ESR-dependent manner. Canavanine-induced mutagenesis also involves translesion DNA polymerases Rev1 and Polζ and non-homologous end joining factor Ku. Furthermore, under conditions of chronic sub-lethal canavanine stress, deletions of Rev1, Polζ, and Ku-encoding genes exhibit genetic interactions with ESR mutants indicative of ESR regulating these mutagenic DNA repair processes. Analyses of mutagenesis induced by several different stresses showed that the ESR specifically modulates mutagenesis induced by proteotoxic stress. Together, these results document the first known example of an involvement of a eukaryotic stress response pathway in mutagenesis and have important implications for mechanisms of evolution, carcinogenesis, and emergence of drug-resistant pathogens and chemotherapy-resistant tumors. PMID:23935537

Higher baseline viral diversity correlates with lower HBsAg decline following PEGylated interferon-alpha therapy in patients with HBeAg-positive chronic hepatitis B.

PubMed

Li, Hu; Zhang, Li; Ren, Hong; Hu, Peng

2018-01-01

Viral diversity seems to predict treatment outcomes in certain viral infections. The aim of this study was to evaluate the association between baseline intra-patient viral diversity and hepatitis B surface antigen (HBsAg) decline following PEGylated interferon-alpha (Peg-IFN-α) therapy. Twenty-six HBeAg-positive patients who were treated with Peg-IFN-α were enrolled. Nested polymerase chain reaction (PCR), cloning, and sequencing of the hepatitis B virus S gene were performed on baseline samples, and normalized Shannon entropy (Sn) was calculated as a measure of small hepatitis B surface protein (SHBs) diversity. Multiple regression analysis was used to estimate the association between baseline Sn and HBsAg decline. Of the 26 patients enrolled in the study, 65.4% were male and 61.5% were infected with hepatitis B virus genotype B. The median HBsAg level at baseline was 4.5 log 10 IU/mL (interquartile range: 4.1-4.9) and declined to 3.0 log 10 IU/mL (interquartile range: 1.7-3.9) after 48 weeks of Peg-IFN-α treatment. In models adjusted for baseline alanine aminotransferase (ALT) and HBsAg, the adjusted coefficients (95% CI) for ΔHBsAg and relative percentage HBsAg decrease were -1.3 (-2.5, -0.2) log 10 IU/mL for higher SHBs diversity (Sn≥0.58) patients and -26.4% (-50.2%, -2.5%) for lower diversity (Sn<0.58) patients. Further analysis showed that the "a" determinant upstream flanking region and the first loop of the "a" determinant (nucleotides 341-359, 371-389, and 381-399) were the main sources of higher SHBs diversity. Baseline intra-patient SHBs diversity was inverse to HBsAg decline in HBeAg-positive chronic hepatitis B (CHB) patients receiving Peg-IFN-α monotherapy. Also, more sequence variations within the "a" determinant upstream flanking region and the first loop of the "a" determinant were the main sources of the higher SHBs diversity.

Cellular stress induces cytoplasmic RNA granules in fission yeast.

PubMed

Nilsson, Daniel; Sunnerhagen, Per

2011-01-01

Severe stress causes plant and animal cells to form large cytoplasmic granules containing RNA and proteins. Here, we demonstrate the existence of stress-induced cytoplasmic RNA granules in Schizosaccharomyces pombe. Homologs to several known protein components of mammalian processing bodies and stress granules are found in fission yeast RNA granules. In contrast to mammalian cells, poly(A)-binding protein (Pabp) colocalizes in stress-induced granules with decapping protein. After glucose deprivation, protein kinase A (PKA) is required for accumulation of Pabp-positive granules and translational down-regulation. This is the first demonstration of a role for PKA in RNA granule formation. In mammals, the translation initiation protein eIF2α is a key regulator of formation of granules containing poly(A)-binding protein. In S. pombe, nonphosphorylatable eIF2α does not block but delays granule formation and subsequent clearance after exposure to hyperosmosis. At least two separate pathways in S. pombe appear to regulate stress-induced granules: pka1 mutants are fully proficient to form granules after hyperosmotic shock; conversely, eIF2α does not affect granule formation in glucose starvation. Further, we demonstrate a Pka1-dependent link between calcium perturbation and RNA granules, which has not been described earlier in any organism.

Rapid depletion of budding yeast proteins by fusion to a heat-inducible degron.

PubMed

Sanchez-Diaz, Alberto; Kanemaki, Masato; Marchesi, Vanessa; Labib, Karim

2004-03-02

One effective way to study the biological function of a protein in vivo is to inactivate it and see what happens to the cell. For proteins that are dispensable for cell viability, the corresponding gene can simply be deleted from its chromosomal locus. The study of essential proteins is more challenging, however, because the function of the protein must be inactivated conditionally. Here, we describe a method that allows the target protein to be depleted rapidly and conditionally, so that the immediate effects on the cell can be examined. The chromosomal locus of a budding yeast gene is modified so that a "heat-inducible degron cassette" is added to the N terminus of the encoded protein, causing it to be degraded by a specific ubiquitin-mediated pathway when cells are shifted from 24 degrees to 37 degrees C. Degradation requires recognition of the degron cassette by the evolutionarily conserved Ubr1 protein, which is associated with a ubiquitin-conjugating enzyme. To promote rapid and conditional depletion of the target protein, we use a yeast strain in which expression of the UBR1 gene can be either repressed or strongly induced. Degron strains are constructed by a simple "one-step" approach using the polymerase chain reaction.

Preprotein transport machineries of yeast mitochondrial outer membrane are not required for Bax-induced release of intermembrane space proteins.

PubMed

Sanjuán Szklarz, Luiza K; Kozjak-Pavlovic, Vera; Vögtle, F-Nora; Chacinska, Agnieszka; Milenkovic, Dusanka; Vogel, Sandra; Dürr, Mark; Westermann, Benedikt; Guiard, Bernard; Martinou, Jean-Claude; Borner, Christoph; Pfanner, Nikolaus; Meisinger, Chris

2007-04-20

The mitochondrial outer membrane contains protein import machineries, the translocase of the outer membrane (TOM) and the sorting and assembly machinery (SAM). It has been speculated that TOM or SAM are required for Bax-induced release of intermembrane space (IMS) proteins; however, experimental evidence has been scarce. We used isolated yeast mitochondria as a model system and report that Bax promoted an efficient release of soluble IMS proteins while preproteins were still imported, excluding an unspecific damage of mitochondria. Removal of import receptors by protease treatment did not inhibit the release of IMS proteins by Bax. Yeast mutants of each Tom receptor and the Tom40 channel were not impaired in Bax-induced protein release. We analyzed a large collection of mutants of mitochondrial outer membrane proteins, including SAM, fusion and fission components, but none of these components was required for Bax-induced protein release. The released proteins included complexes up to a size of 230 kDa. We conclude that Bax promotes efficient release of IMS proteins through the outer membrane of yeast mitochondria while the inner membrane remains intact. Inactivation of the known protein import and sorting machineries of the outer membrane does not impair the function of Bax at the mitochondria.

Selection and Characterization of Potential Baker's Yeast from Indigenous Resources of Nepal

PubMed Central

Timilsina, Parash Mani; Yadav, Archana; Joshi, Yogesh; Bhujel, Sahansila; Adhikari, Rojina; Neupane, Katyayanee

2017-01-01

The study aims to isolate the yeast strains that could be used effectively as baker's yeast and compare them with the commercial baker's yeast available in the market of Nepal. A total of 10 samples including locally available sources like fruits, Murcha, and a local tree “Dar” were collected from different localities of Bhaktapur, Kavre, and Syangja districts of Nepal, respectively. Following enrichment and fermentation of the samples, 26 yeast strains were isolated using selective medium Wallerstein Laboratory Nutrient Agar. From the differential tests which included morphological and microscopic observation and physiological and biochemical characterization such as nitrate reduction and lactose utilization tests, 8 strains were selected as possible Saccharomyces strain. The selected strains were further assessed for their efficient leavening ability by tests such as ethanol tolerance, osmotolerance, invertase test, and stress exclusion test. The three most potent strains ENG, MUR3B, and SUG1 isolated from grape, Murcha, and sugarcane, respectively, were used in the fermentation and baking of dough. These strains also carried a possibility of being used as industrial baker's yeast. PMID:29387490

Selection and Characterization of Potential Baker's Yeast from Indigenous Resources of Nepal.

PubMed

Karki, Tika B; Timilsina, Parash Mani; Yadav, Archana; Pandey, Gyanu Raj; Joshi, Yogesh; Bhujel, Sahansila; Adhikari, Rojina; Neupane, Katyayanee

2017-01-01

The study aims to isolate the yeast strains that could be used effectively as baker's yeast and compare them with the commercial baker's yeast available in the market of Nepal. A total of 10 samples including locally available sources like fruits, Murcha, and a local tree "Dar" were collected from different localities of Bhaktapur, Kavre, and Syangja districts of Nepal, respectively. Following enrichment and fermentation of the samples, 26 yeast strains were isolated using selective medium Wallerstein Laboratory Nutrient Agar. From the differential tests which included morphological and microscopic observation and physiological and biochemical characterization such as nitrate reduction and lactose utilization tests, 8 strains were selected as possible Saccharomyces strain. The selected strains were further assessed for their efficient leavening ability by tests such as ethanol tolerance, osmotolerance, invertase test, and stress exclusion test. The three most potent strains ENG, MUR3B, and SUG1 isolated from grape, Murcha, and sugarcane, respectively, were used in the fermentation and baking of dough. These strains also carried a possibility of being used as industrial baker's yeast.

Eighteen new oleaginous yeast species.

PubMed

Garay, Luis A; Sitepu, Irnayuli R; Cajka, Tomas; Chandra, Idelia; Shi, Sandy; Lin, Ting; German, J Bruce; Fiehn, Oliver; Boundy-Mills, Kyria L

2016-07-01

Of 1600 known species of yeasts, about 70 are known to be oleaginous, defined as being able to accumulate over 20 % intracellular lipids. These yeasts have value for fundamental and applied research. A survey of yeasts from the Phaff Yeast Culture Collection, University of California Davis was performed to identify additional oleaginous species within the Basidiomycota phylum. Fifty-nine strains belonging to 34 species were grown in lipid inducing media, and total cell mass, lipid yield and triacylglycerol profiles were determined. Thirty-two species accumulated at least 20 % lipid and 25 species accumulated over 40 % lipid by dry weight. Eighteen of these species were not previously reported to be oleaginous. Triacylglycerol profiles were suitable for biodiesel production. These results greatly expand the number of known oleaginous yeast species, and reveal the wealth of natural diversity of triacylglycerol profiles within wild-type oleaginous Basidiomycetes.

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

Dexamethasone potently enhances phorbol ester-induced IL-1beta gene expression and nuclear factor NF-kappaB activation.

PubMed

Wang, Y; Zhang, J J; Dai, W; Lei, K Y; Pike, J W

1997-07-15

The synthetic glucocorticoid dexamethasone, an immunosuppressive and anti-inflammatory agent, was investigated for its effect on PMA-mediated expression of the inflammatory cytokine IL-1beta in the human monocytic leukemic cell line THP-1. PMA alone induced the production of low levels of IL-1beta in THP-1 cells, whereas dexamethasone alone had no effect. However, dexamethasone potently enhanced PMA-mediated IL-1beta production. Using a selective and potent inhibitor of protein kinase C, we found that synergistic interaction between PMA and dexamethasone requires protein kinase C activation. PMA has been known to activate nuclear factor NF-kappaB in THP-1 cells. Using an oligonucleotide probe corresponding to an NF-kappaB DNA-binding motif of the IL-1beta gene promoter in gel electrophoresis mobility shift assays, we demonstrated that PMA-induced NF-kappaB activation was greatly potentiated by dexamethasone. Our results indicate that glucocorticoids can be positive regulators of inflammatory cytokine gene expression during monocytic cell differentiation.

Forces in yeast flocculation

NASA Astrophysics Data System (ADS)

El-Kirat-Chatel, Sofiane; Beaussart, Audrey; Vincent, Stéphane P.; Abellán Flos, Marta; Hols, Pascal; Lipke, Peter N.; Dufrêne, Yves F.

2015-01-01

In the baker's yeast Saccharomyces cerevisiae, cell-cell adhesion (``flocculation'') is conferred by a family of lectin-like proteins known as the flocculin (Flo) proteins. Knowledge of the adhesive and mechanical properties of flocculins is important for understanding the mechanisms of yeast adhesion, and may help controlling yeast behaviour in biotechnology. We use single-molecule and single-cell atomic force microscopy (AFM) to explore the nanoscale forces engaged in yeast flocculation, focusing on the role of Flo1 as a prototype of flocculins. Using AFM tips labelled with mannose, we detect single flocculins on Flo1-expressing cells, showing they are widely exposed on the cell surface. When subjected to force, individual Flo1 proteins display two distinct force responses, i.e. weak lectin binding forces and strong unfolding forces reflecting the force-induced extension of hydrophobic tandem repeats. We demonstrate that cell-cell adhesion bonds also involve multiple weak lectin interactions together with strong unfolding forces, both associated with Flo1 molecules. Single-molecule and single-cell data correlate with microscale cell adhesion behaviour, suggesting strongly that Flo1 mechanics is critical for yeast flocculation. These results favour a model in which not only weak lectin-sugar interactions are involved in yeast flocculation but also strong hydrophobic interactions resulting from protein unfolding.

The requirement of sterol glucoside for pexophagy in yeast is dependent on the species and nature of peroxisome inducers.

PubMed

Nazarko, Taras Y; Polupanov, Andriy S; Manjithaya, Ravi R; Subramani, Suresh; Sibirny, Andriy A

2007-01-01

Sterol glucosyltransferase, Ugt51/Atg26, is essential for both micropexophagy and macropexophagy of methanol-induced peroxisomes in Pichia pastoris. However, the role of this protein in pexophagy in other yeast remained unclear. We show that oleate- and amine-induced peroxisomes in Yarrowia lipolytica are degraded by Atg26-independent macropexophagy. Surprisingly, Atg26 was also not essential for macropexophagy of oleate- and amine-induced peroxisomes in P. pastoris, suggesting that the function of sterol glucoside (SG) in pexophagy is both species and peroxisome inducer specific. However, the rates of degradation of oleate- and amine-induced peroxisomes in P. pastoris were reduced in the absence of SG, indicating that P. pastoris specifically uses sterol conversion by Atg26 to enhance selective degradation of peroxisomes. However, methanol-induced peroxisomes apparently have lost the redundant ability to be degraded without SG. We also show that the P. pastoris Vac8 armadillo repeat protein is not essential for macropexophagy of methanol-, oleate-, or amine-induced peroxisomes, which makes PpVac8 the first known protein required for the micropexophagy, but not for the macropexophagy, machinery. The uniqueness of Atg26 and Vac8 functions under different pexophagy conditions demonstrates that not only pexophagy inducers, such as glucose or ethanol, but also the inducers of peroxisomes, such as methanol, oleate, or primary amines, determine the requirements for subsequent pexophagy in yeast.

Forsythoside A exerts antipyretic effect on yeast-induced pyrexia mice via inhibiting transient receptor potential vanilloid 1 function

PubMed Central

Liu, Cuiling; Su, Hongchang; Wan, Hongye; Qin, Qingxia; Wu, Xuan; Kong, Xiangying; Lin, Na

2017-01-01

Transient receptor potential vanilloid 1 (TRPV1) is a non-selective cation channel gated by noxious heat, playing major roles in thermoregulation. Forsythoside A (FT-A) is the most abundant phenylethanoid glycosides in Fructus Forsythiae, which has been prescribed as a medicinal herb for treating fever in China for a long history. However, how FT-A affects pyrexia and what is the underlying molecular mechanism remain largely unknown. Here we found that FT-A exerted apparent antipyretic effect through decreasing the levels of prostaglandin E2 (PGE2) and interleukin 8 (IL-8) in a dose-dependent fashion on the yeast induced pyrexia mice. Interestingly, FT-A significantly downregulated TRPV1 expression in the hypothalamus and dorsal root ganglion (DRG) of the yeast induced pyrexia mice. Moreover, FT-A inhibited IL-8 and PGE2 secretions, and calcium influx in the HEK 293T-TRPV1 cells after stimulated with capsaicin, the specific TRPV1 agonist. Further investigation of the molecular mechanisms revealed that FT-A treatment rapidly inhibited phosphorylation of extracellular signal-regulated kinase (ERK), Jun N-terminal kinase (JNK) and p38 in both yeast induced pyrexia mice and HEK 293T-TRPV1 cells. These results suggest that FT-A may serve as a potential antipyretic agent and the therapeutic action of Fructus Forsythiae on pyretic related disease is, in part, due to the FT-A activities. PMID:28123347

Kluyveromyces marxianus and Saccharomyces boulardii Induce Distinct Levels of Dendritic Cell Cytokine Secretion and Significantly Different T Cell Responses In Vitro

PubMed Central

Smith, Ida M.; Baker, Adam; Christensen, Jeffrey E.; Boekhout, Teun; Frøkiær, Hanne; Arneborg, Nils; Jespersen, Lene

2016-01-01

Interactions between members of the intestinal microbiota and the mucosal immune system can significantly impact human health, and in this context, fungi and food-related yeasts are known to influence intestinal inflammation through direct interactions with specialized immune cells in vivo. The aim of the present study was to characterize the immune modulating properties of the food-related yeast Kluyveromyces marxianus in terms of adaptive immune responses indicating inflammation versus tolerance and to explore the mechanisms behind the observed responses. Benchmarking against a Saccharomyces boulardii strain with probiotic effects documented in clinical trials, we evaluated the ability of K. marxianus to modulate human dendritic cell (DC) function in vitro. Further, we assessed yeast induced DC modulation of naive T cells toward effector responses dominated by secretion of IFNγ and IL-17 versus induction of a Treg response characterized by robust IL-10 secretion. In addition, we blocked relevant DC surface receptors and investigated the stimulating properties of β-glucan containing yeast cell wall extracts. K. marxianus and S. boulardii induced distinct levels of DC cytokine secretion, primarily driven by Dectin-1 recognition of β-glucan components in their cell walls. Upon co-incubation of yeast exposed DCs and naive T cells, S. boulardii induced a potent IFNγ response indicating TH1 mobilization. In contrast, K. marxianus induced a response dominated by Foxp3+ Treg cells, a characteristic that may benefit human health in conditions characterized by excessive inflammation and positions K. marxianus as a strong candidate for further development as a novel yeast probiotic. PMID:27898740

Kluyveromyces marxianus and Saccharomyces boulardii Induce Distinct Levels of Dendritic Cell Cytokine Secretion and Significantly Different T Cell Responses In Vitro.

PubMed

Smith, Ida M; Baker, Adam; Christensen, Jeffrey E; Boekhout, Teun; Frøkiær, Hanne; Arneborg, Nils; Jespersen, Lene

2016-01-01

Interactions between members of the intestinal microbiota and the mucosal immune system can significantly impact human health, and in this context, fungi and food-related yeasts are known to influence intestinal inflammation through direct interactions with specialized immune cells in vivo. The aim of the present study was to characterize the immune modulating properties of the food-related yeast Kluyveromyces marxianus in terms of adaptive immune responses indicating inflammation versus tolerance and to explore the mechanisms behind the observed responses. Benchmarking against a Saccharomyces boulardii strain with probiotic effects documented in clinical trials, we evaluated the ability of K. marxianus to modulate human dendritic cell (DC) function in vitro. Further, we assessed yeast induced DC modulation of naive T cells toward effector responses dominated by secretion of IFNγ and IL-17 versus induction of a Treg response characterized by robust IL-10 secretion. In addition, we blocked relevant DC surface receptors and investigated the stimulating properties of β-glucan containing yeast cell wall extracts. K. marxianus and S. boulardii induced distinct levels of DC cytokine secretion, primarily driven by Dectin-1 recognition of β-glucan components in their cell walls. Upon co-incubation of yeast exposed DCs and naive T cells, S. boulardii induced a potent IFNγ response indicating TH1 mobilization. In contrast, K. marxianus induced a response dominated by Foxp3+ Treg cells, a characteristic that may benefit human health in conditions characterized by excessive inflammation and positions K. marxianus as a strong candidate for further development as a novel yeast probiotic.

Identification of Yju3p as functional orthologue of mammalian monoglyceride lipase in the yeast Saccharomycescerevisiae.

PubMed

Heier, Christoph; Taschler, Ulrike; Rengachari, Srinivasan; Oberer, Monika; Wolinski, Heimo; Natter, Klaus; Kohlwein, Sepp D; Leber, Regina; Zimmermann, Robert

2010-09-01

Monoacylglycerols (MAGs) are short-lived intermediates of glycerolipid metabolism. Specific molecular species, such as 2-arachidonoylglycerol, which is a potent activator of cannabinoid receptors, may also function as lipid signaling molecules. In mammals, enzymes hydrolyzing MAG to glycerol and fatty acids, resembling the final step in lipolysis, or esterifying MAG to diacylglycerol, are well known; however, despite the high level of conservation of lipolysis, the corresponding activities in yeast have not been characterized yet. Here we provide evidence that the protein Yju3p functions as a potent MAG hydrolase in yeast. Cellular MAG hydrolase activity was decreased by more than 90% in extracts of Yju3p-deficient cells, indicating that Yju3p accounts for the vast majority of this activity in yeast. Loss of this activity was restored by heterologous expression of murine monoglyceride lipase (MGL). Since yju3Delta mutants accumulated MAG in vivo only at very low concentrations, we considered the possibility that MAGs are re-esterified into DAG by acyltransferases. Indeed, cellular MAG levels were further increased in mutant cells lacking Yju3p and Dga1p or Lro1p acyltransferase activities. In conclusion, our studies suggest that catabolic and anabolic reactions affect cellular MAG levels. Yju3p is the functional orthologue of mammalian MGL and is required for efficient degradation of MAG in yeast. 2010 Elsevier B.V. All rights reserved.

Identification of Yju3p as functional orthologue of mammalian monoglyceride lipase in the yeast Saccharomycescerevisiae

PubMed Central

Heier, Christoph; Taschler, Ulrike; Rengachari, Srinivasan; Oberer, Monika; Wolinski, Heimo; Natter, Klaus; Kohlwein, Sepp D.; Leber, Regina; Zimmermann, Robert

2010-01-01

Monoacylglycerols (MAGs) are short-lived intermediates of glycerolipid metabolism. Specific molecular species, such as 2-arachidonoylglycerol, which is a potent activator of cannabinoid receptors, may also function as lipid signaling molecules. In mammals, enzymes hydrolyzing MAG to glycerol and fatty acids, resembling the final step in lipolysis, or esterifying MAG to diacylglycerol, are well known; however, despite the high level of conservation of lipolysis, the corresponding activities in yeast have not been characterized yet. Here we provide evidence that the protein Yju3p functions as a potent MAG hydrolase in yeast. Cellular MAG hydrolase activity was decreased by more than 90% in extracts of Yju3p-deficient cells, indicating that Yju3p accounts for the vast majority of this activity in yeast. Loss of this activity was restored by heterologous expression of murine monoglyceride lipase (MGL). Since yju3Δ mutants accumulated MAG in vivo only at very low concentrations, we considered the possibility that MAGs are re-esterified into DAG by acyltransferases. Indeed, cellular MAG levels were further increased in mutant cells lacking Yju3p and Dga1p or Lro1p acyltransferase activities. In conclusion, our studies suggest that catabolic and anabolic reactions affect cellular MAG levels. Yju3p is the functional orthologue of mammalian MGL and is required for efficient degradation of MAG in yeast. PMID:20554061

Damage-induced ectopic recombination in the yeast Saccharomyces cerevisiae.

PubMed

Kupiec, M; Steinlauf, R

1997-06-09

Mitotic recombination in the yeast Saccharomyces cerevisiae is induced when cells are irradiated with UV or X-rays, reflecting the efficient repair of damage by recombinational repair mechanisms. We have used multiply marked haploid strains that allow the simultaneous detection of several types of ectopic recombination events. We show that inter-chromosomal ectopic conversion of lys2 heteroalleles and, to a lesser extent, direct repeat recombination (DRR) between non-tandem repeats, are increased by DNA-damaging agents; in contrast, ectopic recombination of the naturally occurring Ty element is not induced. We have tested several hypotheses that could explain the preferential lack of induction of Ty recombination by DNA-damaging agents. We have found that the lack of induction cannot be explained by a cell cycle control or by an effect of the mating-type genes. We also found no role for the flanking long terminal repeats (LTRs) of the Ty in preventing the induction. Ectopic conversion, DRR, and forward mutation of artificial repeats show different kinetics of induction at various positions of the cell cycle, reflecting different mechanisms of recombination. We discuss the mechanistic and evolutionary aspects of these results.

Molecular Characterization of Hap Complex Components Responsible for Methanol-Inducible Gene Expression in the Methylotrophic Yeast Candida boidinii

PubMed Central

Oda, Saori; Yurimoto, Hiroya; Nitta, Nobuhisa; Sasano, Yu

2015-01-01

We identified genes encoding components of the Hap complex, CbHAP2, CbHAP3, and CbHAP5, as transcription factors regulating methanol-inducible gene expression in the methylotrophic yeast Candida boidinii. We found that the Cbhap2Δ, Cbhap3Δ, and Cbhap5Δ gene-disrupted strains showed severe growth defects on methanol but not on glucose and nonfermentable carbon sources such as ethanol and glycerol. In these disruptants, the transcriptional activities of methanol-inducible promoters were significantly decreased compared to those of the wild-type strain, indicating that CbHap2p, CbHap3p, and CbHap5p play indispensable roles in methanol-inducible gene expression. Further molecular and biochemical analyses demonstrated that CbHap2p, CbHap3p, and CbHap5p localized to the nucleus and bound to the promoter regions of methanol-inducible genes regardless of the carbon source, and heterotrimer formation was suggested to be necessary for binding to DNA. Unexpectedly, distinct from Saccharomyces cerevisiae, the Hap complex functioned in methanol-specific induction rather than glucose derepression in C. boidinii. Our results shed light on a novel function of the Hap complex in methanol-inducible gene expression in methylotrophic yeasts. PMID:25595445

[Control levels of Sin3 histone deacetylase for spontaneous and UV-induced mutagenesis in yeasts Saccharomyces cerevisiae].

PubMed

Lebovka, I Iu; Kozhina, T N; Fedorova, I V; Peshekhonov, V T; Evstiukhina, T A; Chernenkov, A Iu; Korolev, V G

2014-01-01

SIN3 gene product operates as a repressor for a huge amount of genes in Saccharomyces cerevisiae. Sin3 protein with a mass of about 175 kDa is a member of the RPD3 protein complex with an assessed mass of greater than 2 million Da. It was previously shownthat RPD3 gene mutations influence recombination and repair processes in S. cerevisiae yeasts. We studied the impacts of the sin3 mutation on UV-light sensitivity and UV-induced mutagenesis in budding yeast cells. The deletion ofthe SIN3 gene causes weak UV-sensitivity of mutant budding cells as compared to the wild-type strain. These results show that the sin3 mutation decreases both spontaneous and UV-induced levels of levels. This fact is hypothetically related to themalfunction of ribonucleotide reductase activity regulation, which leads to a decrease in the dNTP pool and the inaccurate error-prone damage bypass postreplication repair pathway, which in turn provokes a reduction in the incidence of mutations.

Protein Folding Activity of the Ribosome is involved in Yeast Prion Propagation

PubMed Central

Blondel, Marc; Soubigou, Flavie; Evrard, Justine; Nguyen, Phu hai; Hasin, Naushaba; Chédin, Stéphane; Gillet, Reynald; Contesse, Marie-Astrid; Friocourt, Gaëlle; Stahl, Guillaume; Jones, Gary W.; Voisset, Cécile

2016-01-01

6AP and GA are potent inhibitors of yeast and mammalian prions and also specific inhibitors of PFAR, the protein-folding activity borne by domain V of the large rRNA of the large subunit of the ribosome. We therefore explored the link between PFAR and yeast prion [PSI+] using both PFAR-enriched mutants and site-directed methylation. We demonstrate that PFAR is involved in propagation and de novo formation of [PSI+]. PFAR and the yeast heat-shock protein Hsp104 partially compensate each other for [PSI+] propagation. Our data also provide insight into new functions for the ribosome in basal thermotolerance and heat-shocked protein refolding. PFAR is thus an evolutionarily conserved cell component implicated in the prion life cycle, and we propose that it could be a potential therapeutic target for human protein misfolding diseases. PMID:27633137

The galactose-induced decrease in phosphate levels leads to toxicity in yeast models of galactosemia.

PubMed

Machado, Caio M; De-Souza, Evandro A; De-Queiroz, Ana Luiza F V; Pimentel, Felipe S A; Silva, Guilherme F S; Gomes, Fabio M; Montero-Lomelí, Mónica; Masuda, Claudio A

2017-06-01

Classic galactosemia is an inborn error of metabolism caused by deleterious mutations in the GALT gene. A number of evidences indicate that the galactose-1-phosphate accumulation observed in patient cells is a cause of toxicity in this disease. Nevertheless, the consequent molecular events caused by the galactose-1-phosphate accumulation remain elusive. Here we show that intracellular inorganic phosphate levels decreased when yeast models of classic galactosemia were exposed to galactose. The decrease in phosphate levels is probably due to the trapping of phosphate in the accumulated galactose-1-phosphate since the deletion of the galactokinase encoding gene GAL1 suppressed this phenotype. Galactose-induced phosphate depletion caused an increase in glycogen content, an expected result since glycogen breakdown by the enzyme glycogen phosphorylase is dependent on inorganic phosphate. Accordingly, an increase in intracellular phosphate levels suppressed the galactose effect on glycogen content and conferred galactose tolerance to yeast models of galactosemia. These results support the hypothesis that the galactose-induced decrease in phosphate levels leads to toxicity in galactosemia and opens new possibilities for the development of better treatments for this disease. Copyright © 2017 Elsevier B.V. All rights reserved.

Immunizations with chimeric hepatitis B virus-like particles to induce potential anti-hepatitis C virus neutralizing antibodies.

PubMed

Vietheer, Patricia T K; Boo, Irene; Drummer, Heidi E; Netter, Hans-Jürgen

2007-01-01

Virus-like particles (VLPs) are highly immunogenic and proven to induce protective immunity. The small surface antigen (HBsAg-S) of hepatitis B virus (HBV) self-assembles into VLPs and its use as a vaccine results in protective antiviral immunity against HBV infections. Chimeric HBsAg-S proteins carrying foreign epitopes allow particle formation and have the ability to induce anti-foreign humoral and cellular immune responses. The insertion of the hypervariable region 1 (HVR1) sequence derived from the envelope protein 2 (E2) of hepatitis C virus (HCV) into the major antigenic site of HBsAg-S ('a'-determinant) resulted in the formation of highly immunogenic VLPs that retained the antigenicity of the inserted HVR1 sequence. BALB/c mice were immunized with chimeric VLPs, which resulted in antisera with anti-HCV activity. The antisera were able to immunoprecipitate native HCV envelope complexes (E1E2) containing homologous or heterologous HVR1 sequences. HCV E1E2 pseudotyped HIV-1 particles (HCVpp) were used to measure entry into HuH-7 target cells in the presence or absence of antisera that were raised against chimeric VLPs. Anti-HVR1 VLP sera interfered with entry of entry-competent HCVpps containing either homologous or heterologous HVR1 sequences. Also, immunizations with chimeric VLPs induced antisurface antigen (HBsAg) antibodies, indicating that HBV-specific antigenicity and immunogenicity of the 'a'-determinant region is retained. A multivalent vaccine against different pathogens based on the HBsAg delivery platform should be possible. We hypothesize that custom design of VLPs with an appropriate set of HCV-neutralizing epitopes will induce antibodies that would serve to decrease the viral load at the initial infecting inoculum.

Rational design and evaluation of HBsAg polymeric nanoparticles as antigen delivery carriers.

PubMed

Dewangan, Hitesh Kumar; Pandey, Tarun; Maurya, Lakshmi; Singh, Sanjay

2018-05-01

The present work is focused on the development and evaluation of single dose sustained-release Hepatitis B surface antigen (HBsAg) loaded nanovaccine for Hepatitis B. The conventional treatment suffers from repeated administration and hence requires a booster dose. Therefore, polymeric nanovaccine of HBsAg was developed by double emulsion solvent evaporation technique, utilizing central composite design for formulation optimization. The effects of independent variables (like polymer amount, stabilizer concentration, aqueous/organic phase ratio and homogenizer speed) were also studied on critical quality attributes like particle size and entrapment efficiency. Nanovaccine was characterized in terms of physicochemical parameters, release, internalization and in vivo immunological evaluation in BALB/c mice after administration by different routes such as oral, sub-cutaneous, nasal and intramuscular. The designed nanovaccine demonstrated nanometric size with smooth surface, negative zeta potential, maximum entrapment, sustained release and better internalization in macrophage and MRC-5 cell line. The immune-stimulating activity of nanovaccine administered by different routes was evaluated by measuring anti-HBsAg titre like specific immunoglobulin IgG and IgA response and cytokine level (interleukin-2, interferon-Y) measurement. The results indicated that the nanovaccine administered by intramuscular route produced better humoral as well as cellular responses and potential carriers for antigen delivery at single dose administration via intramuscular route. Copyright © 2018 Elsevier B.V. All rights reserved.

Discovery of 4-anilino-N-methylthieno[3,2-d]pyrimidines and 4-anilino-N-methylthieno[2,3-d]pyrimidines as potent apoptosis inducers.

PubMed

Kemnitzer, William; Sirisoma, Nilantha; May, Chris; Tseng, Ben; Drewe, John; Cai, Sui Xiong

2009-07-01

We report the discovery of N-((benzo[d][1,3]dioxol-5-yl)methyl)-6-phenylthieno[3,2-d]pyrimidin-4-amine (2a) as an apoptosis inducer using our proprietary cell- and caspase-based ASAP HTS assay, and SAR study of HTS hit 2a which led to the discovery of 4-anilino-N-methylthieno[3,2-d]pyrimidines and 4-anilino-N-methylthieno[2,3-d]pyrimidines as potent apoptosis inducers. Compounds 5d and 5e were the most potent with EC(50) values of 0.008 and 0.004microM in T47D human breast cancer cells, respectively. Compound 5d was found to be highly active in the MX-1 breast cancer model. Functionally, compounds 5d and 5e both induced apoptosis through inhibition of tubulin polymerization.

Induced acute ruminal acidosis in goats treated with yeast (Saccharomyces cerevisiae) and bicarbonate.

PubMed

Aslan, V; Thamsborg, S M; Jørgensen, R J; Basse, A

1995-01-01

Ruminal acidosis was induced in twenty-one 10-month-old West African Dwarf Goats by feeding a suspension of 80 g wheat flour per kg bodyweight (day 0) through a stomach tube. Ruminal and systemic acidosis was diagnosed on day 1 in all goats. Clinical signs included loss of rumination and appetite, trembling, and watery diarrhoea. The detection of acidic faeces during the first 24h was considered of diagnostic importance. Subgroups were treated orally on days 1, 2, and 3 either with 1 g of sodium bicarbonate per kg bodyweight, with 1 g of baking yeast per kg, or with a combination of these treatments at 0.5 g of each per kg. A fourth group served as untreated controls. Peroral bicarbonate neutralization was highly effective in the treatment of rumen acidosis, whereas the use of yeast was found ineffective. The combined treatment had a moderate effect probably due to the bicarbonate. Three fatal cases (60%) occurred in the untreated group compared with none in the bicarbonate group, and 2 in each of the remaining groups. This corresponded to 33% of the yeast treated group and 40% of the combined treated group. Details were given on post mortem examinations performed on all survivors on day 11. Lesions included subacute rumenitis and abomasal ulcers. No lesions were found in 3 of the bicarbonate treated goats and in 2 of the animals receiving combined treatment.

Micropropagation of transgenic lettuce containing HBsAg as a method of mass-scale production of standardised plant material for biofarming purposes.

PubMed

Pniewski, Tomasz; Czyż, Marcin; Wyrwa, Katarzyna; Bociąg, Piotr; Krajewski, Paweł; Kapusta, Józef

2017-01-01

Micropropagation protocol of transgenic lettuce bearing S-, M- and L-HBsAg was developed for increased production of uniformised material for oral vaccine preparation. Effective manufacturing of plant-based biopharmaceuticals, including oral vaccines, depends on sufficient content of a protein of interest in the initial material and its efficient conversion into an administrable formulation. However, stable production of plants with a uniformised antigen content is equally important for reproducible processing. This can be provided by micropropagation techniques. Here, we present a protocol for micropropagation of transgenic lettuce lines bearing HBV surface antigens: S-, M- and L-HBsAg. These were multiplied through axillary buds to avoid the risk of somaclonal variation. Micropropagation effectiveness reached 3.5-5.7 per passage, which implies potential production of up to 6600 plant clones within a maximum 5 months. Multiplication and rooting rates were statistically homogenous for most transgenic and control plants. For most lines, more than 90 % of clones obtained via in vitro micropropagation had HBsAg content as high as reference plants directly developed from seeds. Clones were also several times more uniform in HBsAg expression. Variation coefficients of HBsAg content did not exceed 10 % for approximately 40-85 % of clones, or reached a maximum 20 % for 90 % of all clones. Tissue culture did not affect total and leaf biomass yields. Seed production for clones was decreased insignificantly and did not impact progeny condition. Micropropagation facilitates a substantial increase in the production of lettuce plants with high and considerably equalised HBsAg contents. This, together with the previously reported optimisation of plant tissue processing and its long-term stability, constitutes a successive step in manufacturing of a standardised anti-HBV oral vaccine of reliable efficacy.

Increased heme synthesis in yeast induces a metabolic switch from fermentation to respiration even under conditions of glucose repression.

PubMed

Zhang, Tiantian; Bu, Pengli; Zeng, Joey; Vancura, Ales

2017-10-13

Regulation of mitochondrial biogenesis and respiration is a complex process that involves several signaling pathways and transcription factors as well as communication between the nuclear and mitochondrial genomes. Under aerobic conditions, the budding yeast Saccharomyces cerevisiae metabolizes glucose predominantly by glycolysis and fermentation. We have recently shown that altered chromatin structure in yeast induces respiration by a mechanism that requires transport and metabolism of pyruvate in mitochondria. However, how pyruvate controls the transcriptional responses underlying the metabolic switch from fermentation to respiration is unknown. Here, we report that this pyruvate effect involves heme. We found that heme induces transcription of HAP4 , the transcriptional activation subunit of the Hap2/3/4/5p complex, required for growth on nonfermentable carbon sources, in a Hap1p- and Hap2/3/4/5p-dependent manner. Increasing cellular heme levels by inactivating ROX1 , which encodes a repressor of many hypoxic genes, or by overexpressing HEM3 or HEM12 induced respiration and elevated ATP levels. Increased heme synthesis, even under conditions of glucose repression, activated Hap1p and the Hap2/3/4/5p complex and induced transcription of HAP4 and genes required for the tricarboxylic acid (TCA) cycle, electron transport chain, and oxidative phosphorylation, leading to a switch from fermentation to respiration. Conversely, inhibiting metabolic flux into the TCA cycle reduced cellular heme levels and HAP4 transcription. Together, our results indicate that the glucose-mediated repression of respiration in budding yeast is at least partly due to the low cellular heme level. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Heteroassembled gold nanoparticles with sandwich-immunoassay LSPR chip format for rapid and sensitive detection of hepatitis B virus surface antigen (HBsAg).

PubMed

Kim, Jinwoon; Oh, Seo Yeong; Shukla, Shruti; Hong, Seok Bok; Heo, Nam Su; Bajpai, Vivek K; Chun, Hyang Sook; Jo, Cheon-Ho; Choi, Bong Gill; Huh, Yun Suk; Han, Young-Kyu

2018-06-01

This study aimed to develop a more sensitive method for the detection of hepatitis B surface antigen (HBsAg) using heteroassembled gold nanoparticles (AuNPs). A single layered localized surface plasmon resonance (LSPR) chip format was developed with antigen-antibody reaction-based detection symmetry using AuNPs, which detected HBsAg at 10 pg/mL. To further improve the detection limit, a modified detection format was fabricated by fixing a secondary antibody (to form a heteroassembled sandwich format) to the AuNP monolayer, which enhanced the detection sensitivity by about 100 times. The developed heteroassembled AuNPs sandwich-immunoassay LSPR chip format was able to detect as little as 100 fg/mL of HBsAg within 10-15 min. In addition, the heteroassembled AuNPs sandwich-immunoassay LSPR chip format did not show any non-specific binding to other tested antigens, including alpha fetoprotein (AFP), C-reactive protein (CRP), and prostate-specific antigen (PSA). These findings confirm that the proposed detection strategy of heteroassembled AuNPs sandwich-immunoassay LSPR chip format may provide a new platform for early diagnosis of various human diseases. Copyright © 2018 Elsevier B.V. All rights reserved.

Expression of hepatitis B surface antigen in transgenic plants.

PubMed Central

Mason, H S; Lam, D M; Arntzen, C J

1992-01-01

Tobacco plants were genetically transformed with the gene encoding hepatitis B surface antigen (HBsAg) linked to a nominally constitutive promoter. Enzyme-linked immunoassays using a monoclonal antibody directed against human serum-derived HBsAg revealed the presence of HBsAg in extracts of transformed leaves at levels that correlated with mRNA abundance. This suggests that there were no major inherent limitations of transcription or translation of this foreign gene in plants. Recombinant HBsAg was purified from transgenic plants by immunoaffinity chromatography and examined by electron microscopy. Spherical particles with an average diameter of 22 nm were observed in negatively stained preparations. Sedimentation of transgenic plant extracts in sucrose and cesium chloride density gradients showed that the recombinant HBsAg and human serum-derived HBsAg had similar physical properties. Because the HBsAg produced in transgenic plants is antigenically and physically similar to the HBsAg particles derived from human serum and recombinant yeast, which are used as vaccines, we conclude that transgenic plants hold promise as low-cost vaccine production systems. Images PMID:1465391

Liposomal short-chain C6 ceramide induces potent anti-osteosarcoma activity in vitro and in vivo.

PubMed

Zhai, Lei; Sun, Nan; Han, Zhe; Jin, Hai-chao; Zhang, Bo

Osteosarcoma (OS) remains one deadly disease for many affected patients. The search for novel and more efficient anti-OS agents is urgent. In the current study, we demonstrated that liposome-packed C6 ceramide exerted potent cytotoxic effect against established (U2OS and MG-63 lines) and primary human OS cells. Meanwhile, the liposomal C6 (ceramide) induced caspase-mediated apoptotic death in OS cells. Liposomal C6 was significantly more potent than conventional free C6 in inhibiting OS cells, yet it was safe to non-cancerous bone cells (primary murine osteoblasts or human MLO-Y4 osteocytic cells). At the signaling level, we showed that liposomal C6 potently inhibited Akt activation in OS cells. Further studies revealed that a low dose of liposomal C6 dramatically sensitized the in vitro anti-OS activity of two conventional chemodrugs: methotrexate (MTX) and doxorubicin. In vivo, intravenous injection of liposomal C6 inhibited Akt activation and suppressed U2OS xenograft growth in nude mice without causing apparent toxicities. Meanwhile, when given at a low-dose (5 mg/kg body weight), liposomal C6 dramatically sensitized MTX's anti-U2OS activity in vivo. Collectively, our data demonstrate that liposomal C6 exerts potent anti-tumor activity in preclinical OS models. Copyright © 2015 Elsevier Inc. All rights reserved.

Inactive and mutagenic effects induced by carbon beams of different LET values in a red yeast strain

NASA Astrophysics Data System (ADS)

Wang, Jufang; Lu, Dong; Wu, Xin; Sun, Haining; Ma, Shuang; Li, Renmin; Li, Wenjian

2010-09-01

To evaluate biological action of microorganism exposed to charged particles during the long distance space exploration, induction of inactivation and mutation in a red yeast strain Rhodotorula glutinis AY 91015 by carbon beams of different LET values (14.9-120.0 keV μm -1) was investigated. It was found that survival curves were exponential, and mutation curves were linear for all LET values. The dependence of inactivation cross section on LET approached saturation near 120.0 keV μm -1. The mutation cross section saturated when LET was higher than 58.2 keV μm -1. Meanwhile, the highest RBE i for inactivation located at 120.0 keV μm -1 and the highest RBE m for mutation was at 58.2 keV μm -1. The experiments imply that the most efficient mutagenic part of the depth dose profile of carbon ion is at the plateau region with intermediate LET value in which energy deposited is high enough to induce mutagenic lesions but too low to induce over kill effect in the yeast cells.

Hormetic concentrations of hydrogen peroxide but not ethanol induce cross-adaptation to different stresses in budding yeast.

PubMed

Semchyshyn, Halyna M

2014-01-01

The biphasic-dose response of microorganisms to hydrogen peroxide is a phenomenon of particular interest in hormesis research. In different animal models, the dose-response curve for ethanol is also nonlinear showing an inhibitory effect at high doses but a stimulatory effect at low doses. In this study, we observed the hormetic-dose response to ethanol in budding yeast S. cerevisiae. Cross-protection is a phenomenon in which exposure to mild stress results in the acquisition of cellular resistance to lethal stress induced by different factors. Since both hydrogen peroxide and ethanol at low concentrations were found to stimulate yeast colony growth, we evaluated the role of one substance in cell cross-adaptation to the other substance as well as some weak organic acid preservatives. This study demonstrates that, unlike ethanol, hydrogen peroxide at hormetic concentrations causes cross-resistance of S. cerevisiae to different stresses. The regulatory protein Yap1 plays an important role in the hormetic effects by low concentrations of either hydrogen peroxide or ethanol, and it is involved in the yeast cross-adaptation by low sublethal doses of hydrogen peroxide.

Carbonation acceleration of calcium hydroxide nanoparticles: induced by yeast fermentation

NASA Astrophysics Data System (ADS)

Lopez-Arce, Paula; Zornoza-Indart, Ainara

2015-09-01

Carbonation of Ca(OH)2 nanoparticles and consolidation of limestone are accelerated by high humidity and a yeast fermentation system that supplies a saturated atmosphere on CO2, H2O vapor and ethanol during 28 days. Nanoparticles were analyzed by X-ray diffraction and differential thermal analyses with thermogravimetry. Spectrophotometry, scanning electron microscopy analyses, and hydric and mechanical tests were also performed in stones specimens. Samples exposed to the yeast environment achieve 100 % relative CaCO3 yield, whereas at high humidity but without the yeast and under laboratory environment, relative yields of 95 % CaCO3 and 15 % CaCO3 are, respectively, reached, with white crusts and glazing left on the stone surfaces when the nanoparticles are applied at a concentration of 25 g/l. The largest increase in the drilling resistance and surface hardness values with slight increase in the capillarity absorption and desorption coefficients and with lesser stone color changes are produced at a concentration of 5 g/l, in the yeast system environment. This especially happens in stone specimens initially with bimodal pore size distributions, more amounts of pores with diameters between 0.1 and 1 µm, higher open porosity values and faster capillary coefficients. An inexpensive and reliable method based on water and yeast-sugar solution is presented to speed up carbonation of Ca(OH)2 nanoparticles used as a consolidating product to improve the mechanical properties of decayed limestone from archaeological and architectural heritage.

HIV-1 Protease in the Fission Yeast Schizosaccharomyces pombe.

PubMed

Benko, Zsigmond; Elder, Robert T; Li, Ge; Liang, Dong; Zhao, Richard Y

2016-01-01

HIV-1 protease (PR) is an essential viral enzyme. Its primary function is to proteolyze the viral Gag-Pol polyprotein for production of viral enzymes and structural proteins and for maturation of infectious viral particles. Increasing evidence suggests that PR cleaves host cellular proteins. However, the nature of PR-host cellular protein interactions is elusive. This study aimed to develop a fission yeast (Schizosaccharomyces pombe) model system and to examine the possible interaction of HIV-1 PR with cellular proteins and its potential impact on cell proliferation and viability. A fission yeast strain RE294 was created that carried a single integrated copy of the PR gene in its chromosome. The PR gene was expressed using an inducible nmt1 promoter so that PR-specific effects could be measured. HIV-1 PR from this system cleaved the same indigenous viral p6/MA protein substrate as it does in natural HIV-1 infections. HIV-1 PR expression in fission yeast cells prevented cell proliferation and induced cellular oxidative stress and changes in mitochondrial morphology that led to cell death. Both these PR activities can be prevented by a PR-specific enzymatic inhibitor, indinavir, suggesting that PR-mediated proteolytic activities and cytotoxic effects resulted from enzymatic activities of HIV-1 PR. Through genome-wide screening, a serine/threonine kinase, Hhp2, was identified that suppresses HIV-1 PR-induced protease cleavage and cell death in fission yeast and in mammalian cells, where it prevented PR-induced apoptosis and cleavage of caspase-3 and caspase-8. This is the first report to show that HIV-1 protease is functional as an enzyme in fission yeast, and that it behaves in a similar manner as it does in HIV-1 infection. HIV-1 PR-induced cell death in fission yeast could potentially be used as an endpoint for mechanistic studies, and this system could be used for developing a high-throughput system for drug screenings.

Break-induced replication and recombinational telomere elongation in yeast.

PubMed

McEachern, Michael J; Haber, James E

2006-01-01

When a telomere becomes unprotected or if only one end of a chromosomal double-strand break succeeds in recombining with a template sequence, DNA can be repaired by a recombination-dependent DNA replication process termed break-induced replication (BIR). In budding yeasts, there are two BIR pathways, one dependent on the Rad51 recombinase protein and one Rad51 independent; these two repair processes lead to different types of survivors in cells lacking the telomerase enzyme that is required for normal telomere maintenance. Recombination at telomeres is triggered by either excessive telomere shortening or disruptions in the function of telomere-binding proteins. Telomere elongation by BIR appears to often occur through a "roll and spread" mechanism. In this process, a telomeric circle produced by recombination at a dysfunctional telomere acts as a template for a rolling circle BIR event to form an elongated telomere. Additional BIR events can then copy the elongated sequence to all other telomeres.

Potent apoptosis-inducing activity of erypoegin K, an isoflavone isolated from Erythrina poeppigiana, against human leukemia HL-60 cells.

PubMed

Hikita, Kiyomi; Hattori, Natsuki; Takeda, Aya; Yamakage, Yuko; Shibata, Rina; Yamada, Saori; Kato, Kuniki; Murata, Tomiyasu; Tanaka, Hitoshi; Kaneda, Norio

2018-01-01

Erypoegin K is an isoflavone isolated from the stem bark of Erythrina poeppigiana. It contains a furan group at the A-ring of the core isoflavone structure and can inhibit the activity of glyoxalase I, an enzyme that catalyzes the detoxification of methylglyoxal (MG), a by-product of glycolysis. In the present study, we found that erypoegin K has a potent cytotoxic effect on human leukemia HL-60 cells. Its cytotoxic effect was much stronger than that of a known glyoxalase I inhibitor S-p-bromobenzylglutathione cyclopentyl diester. Conversely, erypoegin K demonstrated weak cytotoxicity toward normal human peripheral lymphocytes. The treatment of HL-60 cells with erypoegin K significantly induced caspase-3 activity, whereas the pretreatment of the cells with caspase-3 inhibitor suppressed erypoegin K-induced cell death. Furthermore, nuclear condensation and apoptotic genome DNA fragmentation were observed in erypoegin K-treated HL-60 cells. These results indicated that the observed cell death was mediated by apoptosis. In addition, the toxic compound MG was highly accumulated in the culture medium of erypoegin K-treated HL-60 cells, suggesting that cell apoptosis was triggered by extracellular MG. The present study showed that erypoegin K has a potent apoptosis-inducing effect on cancerous cell lines, such as HL-60.

Cofilin, But Not Profilin, Is Required for Myosin-I-Induced Actin Polymerization and the Endocytic Uptake in Yeast

PubMed Central

Idrissi, Fatima-Zahra; Wolf, Bianka L.; Geli, M. Isabel

2002-01-01

Mutations in the budding yeast myosins-I (MYO3 and MYO5) cause defects in the actin cytoskeleton and in the endocytic uptake. Robust evidence also indicates that these proteins induce Arp2/3-dependent actin polymerization. Consistently, we have recently demonstrated, using fluorescence microscopy, that Myo5p is able to induce cytosol-dependent actin polymerization on the surface of Sepharose beads. Strikingly, we now observed that, at short incubation times, Myo5p induced the formation of actin foci that resembled the yeast cortical actin patches, a plasma membrane-associated structure that might be involved in the endocytic uptake. Analysis of the machinery required for the formation of the Myo5p-induced actin patches in vitro demonstrated that the Arp2/3 complex was necessary but not sufficient in the assay. In addition, we found that cofilin was directly involved in the process. Strikingly though, the cofilin requirement seemed to be independent of its ability to disassemble actin filaments and profilin, a protein that closely cooperates with cofilin to maintain a rapid actin filament turnover, was not needed in the assay. In agreement with these observations, we found that like the Arp2/3 complex and the myosins-I, cofilin was essential for the endocytic uptake in vivo, whereas profilin was dispensable. PMID:12429847

The marine cytotoxin portimine is a potent and selective inducer of apoptosis.

PubMed

Cuddihy, Sarah L; Drake, Sarah; Harwood, D Tim; Selwood, Andrew I; McNabb, Paul S; Hampton, Mark B

2016-12-01

Portimine is a recently discovered member of a class of marine micro-algal toxins called cyclic imines. In dramatic contrast to related compounds in this toxin class, portimine has very low acute toxicity to mice but is highly cytotoxic to cultured cells. In this study we show that portimine kills human Jurkat T-lymphoma cells and mouse embryonic fibroblasts (MEFs), with LC 50 values of 6 and 2.5 nM respectively. Treated cells displayed rapid caspase activation and phosphatidylserine exposure, indicative of apoptotic cell death. Jurkat cells overexpressing the anti-apoptotic protein Bcl-2 or Bax/Bak knockout MEFs were completely protected from portimine. This protection was apparent even at high concentrations of portimine, with no evidence of necrotic cell death, indicating that portimine is a selective chemical inducer of apoptosis. Treatment of the Bcl-2-overexpressing cells with both portimine and the Bcl-2 inhibitor ABT-737 proved a powerful combination, causing >90 % death. We conclude that portimine is one of the most potent naturally derived inducers of apoptosis to be discovered, and it displays strong selectivity for the induction of apoptotic pathways.

Light-mediated control of DNA transcription in yeast

PubMed Central

Hughes, Robert M.; Bolger, Steven; Tapadia, Hersh; Tucker, Chandra L.

2012-01-01

A variety of methods exist for inducible control of DNA transcription in yeast. These include the use of native yeast promoters or regulatory elements that are responsive to small molecules such as galactose, methionine, and copper, or engineered systems that allow regulation by orthogonal small molecules such as estrogen. While chemically regulated systems are easy to use and can yield high levels of protein expression, they often provide imprecise control over protein levels. Moreover, chemically regulated systems can affect many other proteins and pathways in yeast, activating signaling pathways or physiological responses. Here, we describe several methods for light mediated control of DNA transcription in vivo in yeast. We describe methodology for using a red light and phytochrome dependent system to induce transcription of genes under GAL1 promoter control, as well as blue light / cryptochrome dependent systems to control transcription of genes under GAL1 promoter or LexA operator control. Light is dose dependent, inexpensive to apply, easily delivered, and does not interfere with cellular pathways, and thus has significant advantages over chemical systems. PMID:22922268

Differential pathways regulating innate and adaptive antitumor immune responses by particulate and soluble yeast-derived β-glucans

PubMed Central

Qi, Chunjian; Cai, Yihua; Gunn, Lacey; Ding, Chuanlin; Li, Bing; Kloecker, Goetz; Qian, Keqing; Vasilakos, John; Saijo, Shinobu; Iwakura, Yoichiro; Yannelli, John R.

2011-01-01

β-glucans have been reported to function as a potent adjuvant to stimulate innate and adaptive immune responses. However, β-glucans from different sources are differential in their structure, conformation, and thus biologic activity. Different preparations of β-glucans, soluble versus particulate, further complicate their mechanism of action. Here we show that yeast-derived particulate β-glucan activated dendritic cells (DCs) and macrophages via a C-type lectin receptor dectin-1 pathway. Activated DCs by particulate β-glucan promoted Th1 and cytotoxic T-lymphocyte priming and differentiation in vitro. Treatment of orally administered yeast-derived particulate β-glucan elicited potent antitumor immune responses and drastically down-regulated immunosuppressive cells, leading to the delayed tumor progression. Deficiency of the dectin-1 receptor completely abrogated particulate β-glucan–mediated antitumor effects. In contrast, yeast-derived soluble β-glucan bound to DCs and macrophages independent of the dectin-1 receptor and did not activate DCs. Soluble β-glucan alone had no therapeutic effect but significantly augmented antitumor monoclonal antibody-mediated therapeutic efficacy via a complement activation pathway but independent of dectin-1 receptor. These findings reveal the importance of different preparations of β-glucans in the adjuvant therapy and allow for the rational design of immunotherapeutic protocols usable in clinical trials. PMID:21531981

Development and evaluation of yeast-based GFP and luciferase reporter assays for chemical-induced genotoxicity and oxidative damage.

PubMed

Suzuki, Hajime; Sakabe, Takahiro; Hirose, Yuu; Eki, Toshihiko

2017-01-01

We aimed to develop the bioassays for genotixicity and/or oxidative damage using the recombinant yeast. A genotoxicity assay was developed using recombinant Saccharomyces cerevisiae strain BY4741 with a green fluorescent protein (GFP) reporter plasmid, driven by the DNA damage-responsive RNR3 promoter. Enhanced fluorescence induction was observed in DNA repair-deficient strains treated with methyl methanesulfonate, but not with hydrogen peroxide. A GFP reporter yeast strain driven by the oxidative stress-responsive TRX2 promoter was newly developed to assess oxidative damage, but fluorescence was poorly induced by oxidants. In place of GFP, yeast strains with luciferase gene reporter plasmids (luc2 and luc2CP, encoding stable and unstable luciferase, respectively) were prepared. Transient induction of luciferase activity was clearly detected only in a TRX2 promoter-driven luc2CP reporter strain within 90 min of oxidant exposure. However, luciferase was strongly induced by hydroxyurea in the RNR3 promoter-driven luc2 and GFP reporter strains over 8 h after the exposure, suggesting that the RNR3 promoter is continuously upregulated by DNA damage, whereas the TRX2 promoter is transiently activated by oxidative agents. Luciferase activity levels were also increased in a TRX2-promoter-driven luc2CP reporter strain treated with tert-butyl hydroperoxide and menadione and weakly induced with diamide and diethyl maleate. Weakly enhanced luciferase activity induction was detected in the sod1Δ, sod2Δ, and rad27Δ strains treated with hydrogen peroxide compared with that in the wild-type strain. In conclusion, tests using GFP and stable luciferase reporters are useful for genotoxicity, and oxidative damage can be clearly detected by assay with an unstable luciferase reporter.

Dense fibrillar collagen is a potent inducer of invadopodia via a specific signaling network

PubMed Central

Swatkoski, Stephen; Matsumoto, Kazue; Campbell, Catherine B.; Petrie, Ryan J.; Dimitriadis, Emilios K.; Li, Xin; Mueller, Susette C.; Bugge, Thomas H.; Gucek, Marjan

2015-01-01

Cell interactions with the extracellular matrix (ECM) can regulate multiple cellular activities and the matrix itself in dynamic, bidirectional processes. One such process is local proteolytic modification of the ECM. Invadopodia of tumor cells are actin-rich proteolytic protrusions that locally degrade matrix molecules and mediate invasion. We report that a novel high-density fibrillar collagen (HDFC) matrix is a potent inducer of invadopodia, both in carcinoma cell lines and in primary human fibroblasts. In carcinoma cells, HDFC matrix induced formation of invadopodia via a specific integrin signaling pathway that did not require growth factors or even altered gene and protein expression. In contrast, phosphoproteomics identified major changes in a complex phosphosignaling network with kindlin2 serine phosphorylation as a key regulatory element. This kindlin2-dependent signal transduction network was required for efficient induction of invadopodia on dense fibrillar collagen and for local degradation of collagen. This novel phosphosignaling mechanism regulates cell surface invadopodia via kindlin2 for local proteolytic remodeling of the ECM. PMID:25646088

Designed, synthetically accessible bryostatin analogues potently induce activation of latent HIV reservoirs in vitro

NASA Astrophysics Data System (ADS)

Dechristopher, Brian A.; Loy, Brian A.; Marsden, Matthew D.; Schrier, Adam J.; Zack, Jerome A.; Wender, Paul A.

2012-09-01

Bryostatin is a unique lead in the development of potentially transformative therapies for cancer, Alzheimer's disease and the eradication of HIV/AIDS. However, the clinical use of bryostatin has been hampered by its limited supply, difficulties in accessing clinically relevant derivatives, and side effects. Here, we address these problems through the step-economical syntheses of seven members of a new family of designed bryostatin analogues using a highly convergent Prins-macrocyclization strategy. We also demonstrate for the first time that such analogues effectively induce latent HIV activation in vitro with potencies similar to or better than bryostatin. Significantly, these analogues are up to 1,000-fold more potent in inducing latent HIV expression than prostratin, the current clinical candidate for latent virus induction. This study provides the first demonstration that designed, synthetically accessible bryostatin analogues could serve as superior candidates for the eradication of HIV/AIDS through induction of latent viral reservoirs in conjunction with current antiretroviral therapy.

The steroidal Na+/K+ ATPase inhibitor 3-[(R)-3-pyrrolidinyl]oxime derivative (3-R-POD) induces potent pro-apoptotic responses in colonic tumor cells.

PubMed

Alkahtani, Saad Hussin

2014-06-01

Recently, potent anticancer actions of the steroidal Na(+)/K(+) ATPase inhibitor 3-[(R)-3-pyrrolidinyl]oxime derivative 3 (3-R-POD) have been reported for multiple cell lines, including prostate and lung cancer cells. In the present study, the anticancer action of 3-R-POD was addressed in colonic tumor cells. Treatment of Caco2 colonic tumor cells with increasing concentrations of 3-R-POD induced potent, dose-dependent inhibition of cell growth as measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. In addition, the APOpercentage apoptosis assay revealed significant pro-apoptotic responses, suggesting that the anticancer activity of this steroidal Na(+)/K(+) ATPase inhibitor in colonic tumors takes places mainly through the induction of strong pro-apoptotic effects. Focussing on the molecular mechanism that may regulate these interactions, 3-R-POD was shown to induce significant early actin re-organization and late Protein Kinase B (AKT) de-phosphorylation. Finally, the 3-R-POD-induced inhibition of cell growth and early actin reorganization in colonic cancer cells remained unchanged when cells were pre-treated with pertussis toxin, thus excluding possible interactions of this inhibitor with G-coupled receptors. These results indicate that 3-R-POD induces potent pro-apoptotic responses in colonic tumor cells governed by actin re-organization and inhibition of AKT pro-survival signaling. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

[Effects of 33% grapefruit extract on the growth of the yeast--like fungi, dermatopytes and moulds].

PubMed

Krajewska-Kułak, E; Lukaszuk, C; Niczyporuk, W

2001-01-01

Grapefruit seed extract was discovered by Jacob Harich an american immunologist in 1980. Assessment of the influence of grapefruit extract on the yeast-like fungi strains--Candida albicans growth. Material used in this investigation was ATCC test Candida albicans strains no 10231, 200 of Candida albicans strains, 5 of Candida sp. strains isolated from patients with candidiasis symptoms from different ontocenosis and 12 of dermatophytes and moulds isolated from patients. The susceptibility of the Candida was determined by serial dilution method. It seems that 33% grapefruit extract exert a potent antifungal activity against the yeast like fungi strains and had low activity against dermatophytes and moulds. Further studies in vitro and in vivo on greater number of the yeast-like fungi strains and other fungi species are needed.

Structure-based design, synthesis, and biological evaluation of withaferin A-analogues as potent apoptotic inducers.

PubMed

Llanos, Gabriel G; Araujo, Liliana M; Jiménez, Ignacio A; Moujir, Laila M; Rodríguez, Jaime; Jiménez, Carlos; Bazzocchi, Isabel L

2017-11-10

Apoptosis inducers represent an attractive approach for the discovery and development of anticancer agents. Herein, we report on the development by molecular fine tuning of a withaferin A-based library of 63 compounds (2-64), 53 of them reported for the first time. Their antiproliferative evaluation on HeLa, A-549 and MCF-7 human tumor cell lines identified fifteen analogues displaying higher activity (IC 50 values ranging 0.3-4.8 μM) than the lead (IC 50 values ranging 1.3-10.1 μM) either in lag or log growth phases. SAR analysis revealed that acylation enhances cytotoxicity, suggesting the hydrophobic moiety contributes to the activity, presumably by increasing affinity and/or cell membrane permeability. Further investigation clearly indicated that compounds 3, 11, 12, and 18 induce apoptosis evidenced by chromatin condensation, phosphatidylserine externalization, and caspase-3 activation effects on HeLa cells. The potent capacity to induce apoptosis with concomitant cell loss in G2/M highlights the potential of 27-benzyl analogue (18) as an apoptotic inducer drug candidate. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Exploring the Ubiquitin-Proteasome Protein Degradation Pathway in Yeast

ERIC Educational Resources Information Center

Will, Tamara J.; McWatters, Melissa K.; McQuade, Kristi L.

2006-01-01

This article describes an undergraduate biochemistry laboratory investigating the ubiquitin-proteasome pathway in yeast. In this exercise, the enzyme beta-galactosidase (beta-gal) is expressed in yeast under the control of a stress response promoter. Following exposure to heat stress to induce beta-gal expression, cycloheximide is added to halt…

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.

Physiological and environmental control of yeast prions

PubMed Central

Chernova, Tatiana A.; Wilkinson, Keith D.; Chernoff, Yury O.

2014-01-01

Prions are self-perpetuating protein isoforms that cause fatal and incurable neurodegenerative disease in mammals. Recent evidence indicates that a majority of human proteins involved in amyloid and neural inclusion disorders possess at least some prion properties. In lower eukaryotes, such as yeast, prions act as epigenetic elements, which increase phenotypic diversity by altering a range of cellular processes. While some yeast prions are clearly pathogenic, it is also postulated that prion formation could be beneficial in variable environmental conditions. Yeast and mammalian prions have similar molecular properties. Crucial cellular factors and conditions influencing prion formation and propagation were uncovered in the yeast models. Stress-related chaperones, protein quality control deposits, degradation pathways and cytoskeletal networks control prion formation and propagation in yeast. Environmental stresses trigger prion formation and loss, supposedly acting via influencing intracellular concentrations of the prion-inducing proteins, and/or by localizing prionogenic proteins to the prion induction sites via heterologous ancillary helpers. Physiological and environmental modulation of yeast prions points to new opportunities for pharmacological intervention and/or prophylactic measures targeting general cellular systems rather than the properties of individual amyloids and prions. PMID:24236638

Growth Inhibition and DNA Damage Induced by X-Phenols in Yeast: A Quantitative Structure–Activity Relationship Study

PubMed Central

2017-01-01

Phenolic compounds and their derivatives are ubiquitous constituents of numerous synthetic and natural chemicals that exist in the environment. Their toxicity is mostly attributed to their hydrophobicity and/or the formation of free radicals. In a continuation of the study of phenolic toxicity in a systematic manner, we have examined the biological responses of Saccharomyces cerevisiae to a series of mostly monosubstituted phenols utilizing a quantitative structure–activity relationship (QSAR) approach. The biological end points included a growth assay that determines the levels of growth inhibition induced by the phenols as well as a yeast deletion (DEL) assay that assesses the ability of X-phenols to induce DNA damage or DNA breaks. The QSAR analysis of cell growth patterns determined by IC50 and IC80 values indicates that toxicity is delineated by a hydrophobic, parabolic model. The DEL assay was then utilized to detect genomic deletions in yeast. The increase in the genotoxicity was enhanced by the electrophilicity of the phenolic substituents that were strong electron donors as well as by minimal hydrophobicity. The electrophilicities are represented by Brown’s sigma plus values that are a variant of the Hammett sigma constants. A few mutant strains of genes involved in DNA repair were separately exposed to 2,6-di-tert-butyl-4-methyl-phenol (BHT) and butylated hydroxy anisole (BHA). They were subsequently screened for growth phenotypes. BHA-induced growth defects in most of the DNA repair null mutant strains, whereas BHT was unresponsive. PMID:29302629

Miltefosine has post-antifungal effect and induces apoptosis in Cryptococcus yeasts.

PubMed

Spadari, Cristina de Castro; Vila, Taissa; Rozental, Sonia; Ishida, Kelly

2018-05-29

Cryptococcus spp. are common opportunistic fungal pathogens, particularly in HIV patients. The approved drug miltefosine (MFS) has potential as an alternative antifungal against cryptococcosis; however, the mechanism of action of MFS in Cryptococcus is poorly understood. Here, we examined the effects of MFS on C. neoformans and C. gattii yeasts (planktonic and biofilm lifestyles), to clarify its mechanism of action. MFS presented inhibitory and fungicidal effects against planktonic Cryptococcus cells, with similar activity against dispersion biofilm cells, while sessile biofilm cells were less sensitive to MFS. Interestingly, MFS had post-antifungal effect on Cryptococcus , with a proliferation delay of up to 8.15 h after short exposure to fungicidal doses. MFS at fungicidal concentrations increased plasma membrane permeability, likely due to direct interaction with ergosterol, as suggested by competition assays with exogenous ergosterol. Moreover, MFS reduced the mitochondrial membrane potential, increased ROS production, and induced DNA fragmentation and condensation, all of which are hallmarks of apoptosis. Transmission electron microscopy analysis showed that MFS-treated yeasts had a reduced mucopolysaccharide capsule (confirmed by morphometry in light microscopy), plasma membrane irregularities, mitochondrial swelling and a less conspicuous cell wall. Our results suggest that MFS increases plasma membrane permeability in Cryptococcus via interaction with ergosterol, and also affects the mitochondrial membrane, eventually leading to apoptosis, in line with its fungicidal activity. These findings confirm the potential of MFS as an antifungal against C. neoformans and C. gattii, and warrants further studies to establish clinical protocols for MFS use against cryptococcosis. Copyright © 2018 American Society for Microbiology.

Caloric Restriction-Induced Extension of Chronological Lifespan Requires Intact Respiration in Budding Yeast.

PubMed

Kwon, Young-Yon; Lee, Sung-Keun; Lee, Cheol-Koo

2017-04-01

Caloric restriction (CR) has been shown to extend lifespan and prevent cellular senescence in various species ranging from yeast to humans. Many effects of CR may contribute to extend lifespan. Specifically, CR prevents oxidative damage from reactive oxygen species (ROS) by enhancing mitochondrial function. In this study, we characterized 33 single electron transport chain (ETC) gene-deletion strains to identify CR-induced chronological lifespan (CLS) extension mechanisms. Interestingly, defects in 17 of these 33 ETC gene-deleted strains showed loss of both respiratory function and CR-induced CLS extension. On the contrary, the other 16 respiration-capable mutants showed increased CLS upon CR along with increased mitochondrial membrane potential (MMP) and intracellular adenosine triphosphate (ATP) levels, with decreased mitochondrial superoxide generation. We measured the same parameters in the 17 non-respiratory mutants upon CR. CR simultaneously increased MMP and mitochondrial superoxide generation without altering intracellular ATP levels. In conclusion, respiration is essential for CLS extension by CR and is important for balancing MMP, ROS, and ATP levels.

[Treatment of oil-manufacturing wastewater by yeast-SBR system].

PubMed

Lü, Wen-zhou; Liu, Ying; Huang, Yi-zhen

2008-04-01

Eight yeast strains were applied to a sequencing batch reactor (SBR) to treat high-strength oil-containing wastewater. The removal performance, yeast cultivation method and key factors affecting the stability of system were discussed. The results show yeast sludge with MLSS of 19 g/L and SVI of 35 mL/g can be obtained in 6 d in an open system without any molds and bacteria inhibitor addition; In 30 d continuous wastewater treatment, COD and oil removal rate achieve 86.8%-96.9% and above 99.5% respectively under the influent conditions of the COD of 9000-23000 mg/L and oil of 4500-16000 mg/L; Short period of pH impact brings reversible effects on the system and the sludge retention time can affect the SVI of the yeast; Absence of nitrogen induces morphology conversion of some yeast cells from single cell to filamentous one and impairs the settling capability of the yeast.

Improving industrial yeast strains: exploiting natural and artificial diversity

PubMed Central

Steensels, Jan; Snoek, Tim; Meersman, Esther; Nicolino, Martina Picca; Voordeckers, Karin; Verstrepen, Kevin J

2014-01-01

Yeasts have been used for thousands of years to make fermented foods and beverages, such as beer, wine, sake, and bread. However, the choice for a particular yeast strain or species for a specific industrial application is often based on historical, rather than scientific grounds. Moreover, new biotechnological yeast applications, such as the production of second-generation biofuels, confront yeast with environments and challenges that differ from those encountered in traditional food fermentations. Together, this implies that there are interesting opportunities to isolate or generate yeast variants that perform better than the currently used strains. Here, we discuss the different strategies of strain selection and improvement available for both conventional and nonconventional yeasts. Exploiting the existing natural diversity and using techniques such as mutagenesis, protoplast fusion, breeding, genome shuffling and directed evolution to generate artificial diversity, or the use of genetic modification strategies to alter traits in a more targeted way, have led to the selection of superior industrial yeasts. Furthermore, recent technological advances allowed the development of high-throughput techniques, such as ‘global transcription machinery engineering’ (gTME), to induce genetic variation, providing a new source of yeast genetic diversity. PMID:24724938

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.

Combined nattokinase with red yeast rice but not nattokinase alone has potent effects on blood lipids in human subjects with hyperlipidemia.

PubMed

Yang, Nae-Cherng; Chou, Chien-Wen; Chen, Chung-Yin; Hwang, Kai-Lin; Yang, Yi-Chueh

2009-01-01

The purpose of this randomized, double-blind, placebo-controlled, parallel comparison study was to evaluate the lipid-lowering effect of orally administrated nattokinase and nattokinase combined with red yeast rice (RYR) extract on blood lipids in patients with hyperlipidemia. A total of 47 patients with hyperlipidemia were assigned to one of three groups: 1. nattokinase-mono formula (50 mg/capsule), 2. combined formula of nattokinase with RYR (300 mg of extract/capsule) and 3. placebo. Subjects received a twice daily dose of two capsules for six months. The mono formula showed no effects on blood lipids until month six, while the combined formula ameliorated all of measured lipids starting from month one. In the combined group significant decreases were found with regard to: triglycerides (TG) by 15%, total cholesterol (TC) by 25%, low-density lipoprotein cholesterol (LDL-C) by 41%, TC/high-density lipoprotein cholesterol (HDL-C) ratio by 29.5%, and increases in HDL-C by 7.5%. These changes were sustained until the end of study. After controlling for baseline levels, only the combined group, but not mono group, showed a significant difference (p<0.0001) in TC, LDL-C and TC/HDL-C ratio when compared with the placebo group. In summary, this study provides long-term efficacy of nattokinase supplementation and shows that the combined formula has relatively more potent effects than the mono formula on lowering of blood lipids, suggesting that combined nattokinase with RYR will be a better neutraceutical for patients with hyperlipidemia than nattokinase alone.

Genotoxicity of diphenyl diselenide in bacteria and yeast.

PubMed

Rosa, Renato Moreira; Sulzbacher, Krisley; Picada, Jaqueline Nascimento; Roesler, Rafael; Saffi, Jenifer; Brendel, Martin; Henriques, João Antonio Pêgas

2004-10-10

Diphenyl diselenide (DPDS) is an electrophilic reagent used in the synthesis of a variety of pharmacologically active organic selenium compounds. This may increase the risk of human exposure to the chemical at the workplace. We have determined its mutagenic potential in the Salmonella/microsome assay and used the yeast Saccharomyces cerevisiae to assay for putative genotoxicity, recombinogenicity and to determine whether DNA damage produced by DPDS is repairable. Only in exponentially growing cultures was DPDS able to induce frameshift mutations in S. typhimurium and haploid yeast and to increase crossing over and gene conversion frequencies in diploid strains of S. cerevisiae. Thus, DPDS presents a behavior similar to that of an intercalating agent. Mutants defective in excision-resynthesis repair (rad3, rad1), in error-prone repair (rad6) and in recombinational repair (rad52) showed higher than WT-sensitivity to DPDS. It appears that this compound is capable of inducing single and/or double strand breaks in DNA. An epistatic interaction was shown between rad3-e5 and rad52-1 mutant alleles, indicating that excision-resynthesis and strand-break repair may possess common steps in the repair of DNA damage induced by DPDS. DPDS was able to enhance the mutagenesis induced by oxidative mutagens in bacteria. N-acetylcysteine, a glutathione biosynthesis precursor, prevented mutagenesis induced by DPDS in yeast. We have shown that DPDS is a weak mutagen which probably generates DNA strand breaks through both its intercalating action and pro-oxidant effect.

The HBsAg Prevalence Among Blood Donors From Eastern Mediterranean and Middle Eastern Countries: A Systematic Review and Meta-Analysis.

PubMed

Babanejad, Mehran; Izadi, Neda; Najafi, Farid; Alavian, Seyed Moayed

2016-03-01

The world health organization (WHO) recommends that all blood donations should be screened for evidence of infections, such as hepatitis B. The present study aimed to determine the prevalence of hepatitis B surface antigen (HBsAg) in blood donors at the eastern Mediterranean region office (EMRO) of the WHO and middle eastern countries. A meta-analysis was carried out based on the results of an electronic literature search of PubMed, Ovid, Scopus, and Google Scholar for articles published from January 1, 2000, to August 31, 2015. In accordance with a significant homogeneity test and a large value of I2, the random effects model was used to aggregate data from the studies and produce the pooled estimates using the "Metan" command. We included 66 eligible studies. The pooled prevalence of HBsAg in blood donors of both EMRO and middle eastern (E and M) countries was 2.03% (95% confidence interval [CI]: 1.79 - 2.26). In addition, the prevalence rates in the EMRO countries was 1.99% (95% CI: 1.84 - 2.14) and 1.62% in the Middle Eastern countries (95% CI: 1.36 - 1.88). The prevalence among blood donors with more than one study was 1.58% in Egypt, 0.58% in Iran, 0.67% in Iraq, 2.84% in Pakistan, 3.02% in Saudi Arabia, 1.68% in Turkey, and 5.05% in Yemen. Based on the WHO classification of hepatitis B virus (HBV) prevalence, the prevalence of HBsAg in blood donors from E and M countries reached an intermediate level. However, there were low prevalence levels in some E and M countries.

Ammonium Is Toxic for Aging Yeast Cells, Inducing Death and Shortening of the Chronological Lifespan

PubMed Central

Santos, Júlia

2012-01-01

Here we show that in aging Saccharomyces cerevisiae (budding yeast) cells, NH4 + induces cell death associated with shortening of chronological life span. This effect is positively correlated with the concentration of NH4 + added to the culture medium and is particularly evident when cells are starved for auxotrophy-complementing amino acids. NH4 +-induced cell death is accompanied by an initial small increase of apoptotic cells followed by extensive necrosis. Autophagy is inhibited by NH4 +, but this does not cause a decrease in cell viability. We propose that the toxic effects of NH4 + are mediated by activation of PKA and TOR and inhibition of Sch9p. Our data show that NH4 + induces cell death in aging cultures through the regulation of evolutionary conserved pathways. They may also provide new insights into longevity regulation in multicellular organisms and increase our understanding of human disorders such as hyperammonemia as well as effects of amino acid deprivation employed as a therapeutic strategy. PMID:22615903

Breeding research on sake yeasts in Japan: history, recent technological advances, and future perspectives.

PubMed

Kitagaki, Hiroshi; Kitamoto, Katsuhiko

2013-01-01

Sake is an alcoholic beverage of Japan, with a tradition lasting more than 1,300 years; it is produced from rice and water by fermenting with the koji mold Aspergillus oryzae and sake yeast Saccharomyces cerevisiae. Breeding research on sake yeasts was originally developed in Japan by incorporating microbiological and genetic research methodologies adopted in other scientific areas. Since the advent of a genetic paradigm, isolation of yeast mutants has been a dominant approach for the breeding of favorable sake yeasts. These sake yeasts include (a) those that do not form foams (produced by isolating a mutant that does not stick to foams, thus decreasing the cost of sake production); (b) those that do not produce urea, which leads to the formation of ethyl carbamate, a possible carcinogen (isolated by positive selection in a canavanine-, arginine-, and ornithine-containing medium); (c) those that produce an increased amount of ethyl caproate, an apple-like flavor (produced by isolating a mutant resistant to cerulenin, an inhibitor of fatty-acid synthesis); and (d) those that produce a decreased amount of pyruvate (produced by isolating a mutant resistant to an inhibitor of mitochondrial transport, thus decreasing the amount of diacetyl). Given that sake yeasts perform sexual reproduction, sporulation and mating are potent approaches for their breeding. Recently, the genome sequences of sake yeasts have been determined and made publicly accessible. By utilizing this information, the quantitative trait loci (QTLs) for the brewing characteristics of sake yeasts have been identified, which paves a way to DNA marker-assisted selection of the mated strains. Genetic engineering technologies for experimental yeast strains have recently been established by academic groups, and these technologies have also been applied to the breeding of sake yeasts. Sake yeasts whose genomes have been modified with these technologies correspond to genetically modified organisms (GMOs

XRN1 Is a Species-Specific Virus Restriction Factor in Yeasts

PubMed Central

Rowley, Paul A.; Ho, Brandon; Bushong, Sarah; Johnson, Arlen; Sawyer, Sara L.

2016-01-01

In eukaryotes, the degradation of cellular mRNAs is accomplished by Xrn1 and the cytoplasmic exosome. Because viral RNAs often lack canonical caps or poly-A tails, they can also be vulnerable to degradation by these host exonucleases. Yeast lack sophisticated mechanisms of innate and adaptive immunity, but do use RNA degradation as an antiviral defense mechanism. We find a highly refined, species-specific relationship between Xrn1p and the “L-A” totiviruses of different Saccharomyces yeast species. We show that the gene XRN1 has evolved rapidly under positive natural selection in Saccharomyces yeast, resulting in high levels of Xrn1p protein sequence divergence from one yeast species to the next. We also show that these sequence differences translate to differential interactions with the L-A virus, where Xrn1p from S. cerevisiae is most efficient at controlling the L-A virus that chronically infects S. cerevisiae, and Xrn1p from S. kudriavzevii is most efficient at controlling the L-A-like virus that we have discovered within S. kudriavzevii. All Xrn1p orthologs are equivalent in their interaction with another virus-like parasite, the Ty1 retrotransposon. Thus, Xrn1p appears to co-evolve with totiviruses to maintain its potent antiviral activity and limit viral propagation in Saccharomyces yeasts. We demonstrate that Xrn1p physically interacts with the Gag protein encoded by the L-A virus, suggesting a host-virus interaction that is more complicated than just Xrn1p-mediated nucleolytic digestion of viral RNAs. PMID:27711183

A novel Smac mimetic APG-1387 demonstrates potent antitumor activity in nasopharyngeal carcinoma cells by inducing apoptosis.

PubMed

Li, Ning; Feng, Lin; Han, Hui-Qiong; Yuan, Jing; Qi, Xue-Kang; Lian, Yi-Fan; Kuang, Bo-Hua; Zhang, Yu-Chen; Deng, Cheng-Cheng; Zhang, Hao-Jiong; Yao, You-Yuan; Xu, Miao; He, Gui-Ping; Zhao, Bing-Chun; Gao, Ling; Feng, Qi-Sheng; Chen, Li-Zhen; Yang, Lu; Yang, Dajun; Zeng, Yi-Xin

2016-10-10

Despite advances in the development of radiation against nasopharyngeal carcinoma (NPC), the management of advanced NPC remains a challenge. Smac mimetics are designed to neutralize inhibitor of apoptosis (IAP) proteins, thus reactivating the apoptotic program in cancer cells. In this study, we investigated the effect of a novel bivalent Smac mimetic APG-1387 in NPC. In vitro, APG-1387 in combination with TNF-α potently decreased NPC cell viability by inducing apoptosis in majority of NPC cell lines. The in vitro antitumor effect was RIPK1-dependent, whereas it was independent on IAPs, USP11, or EBV. Of note, the inhibition of NF-κB or AKT pathway rendered resistant NPC cells responsive to the treatment of APG-1387/TNF-α. In vivo, APG-1387 displayed antitumor activity as a single agent at well-tolerated doses, even in an in vitro resistant cell line. In summary, our results demonstrate that APG-1387 exerts a potent antitumor effect on NPC. These findings support clinical evaluation of APG-1387 as a potential treatment for advanced NPC. Copyright © 2016. Published by Elsevier Ireland Ltd.

Modeling Yeast Cell Polarization Induced by Pheromone Gradients

NASA Astrophysics Data System (ADS)

Yi, Tau-Mu; Chen, Shanqin; Chou, Ching-Shan; Nie, Qing

2007-07-01

Yeast cells respond to spatial gradients of mating pheromones by polarizing and projecting up the gradient toward the source. It is thought that they employ a spatial sensing mechanism in which the cell compares the concentration of pheromone at different points on the cell surface and determines the maximum point, where the projection forms. Here we constructed the first spatial mathematical model of the yeast pheromone response that describes the dynamics of the heterotrimeric and Cdc42p G-protein cycles, which are linked in a cascade. Two key performance objectives of this system are (1) amplification—converting a shallow external gradient of ligand to a steep internal gradient of protein components and (2) tracking—following changes in gradient direction. We used simulations to investigate amplification mechanisms that allow tracking. We identified specific strategies for regulating the spatial dynamics of the protein components (i.e. their changing location in the cell) that would enable the cell to achieve both objectives.

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.

Immune-escape mutations and stop-codons in HBsAg develop in a large proportion of patients with chronic HBV infection exposed to anti-HBV drugs in Europe.

PubMed

Colagrossi, Luna; Hermans, Lucas E; Salpini, Romina; Di Carlo, Domenico; Pas, Suzan D; Alvarez, Marta; Ben-Ari, Ziv; Boland, Greet; Bruzzone, Bianca; Coppola, Nicola; Seguin-Devaux, Carole; Dyda, Tomasz; Garcia, Federico; Kaiser, Rolf; Köse, Sukran; Krarup, Henrik; Lazarevic, Ivana; Lunar, Maja M; Maylin, Sarah; Micheli, Valeria; Mor, Orna; Paraschiv, Simona; Paraskevis, Dimitros; Poljak, Mario; Puchhammer-Stöckl, Elisabeth; Simon, François; Stanojevic, Maja; Stene-Johansen, Kathrine; Tihic, Nijaz; Trimoulet, Pascale; Verheyen, Jens; Vince, Adriana; Lepej, Snjezana Zidovec; Weis, Nina; Yalcinkaya, Tülay; Boucher, Charles A B; Wensing, Annemarie M J; Perno, Carlo F; Svicher, Valentina

2018-06-01

HBsAg immune-escape mutations can favor HBV-transmission also in vaccinated individuals, promote immunosuppression-driven HBV-reactivation, and increase fitness of drug-resistant strains. Stop-codons can enhance HBV oncogenic-properties. Furthermore, as a consequence of the overlapping structure of HBV genome, some immune-escape mutations or stop-codons in HBsAg can derive from drug-resistance mutations in RT. This study is aimed at gaining insight in prevalence and characteristics of immune-associated escape mutations, and stop-codons in HBsAg in chronically HBV-infected patients experiencing nucleos(t)ide analogues (NA) in Europe. This study analyzed 828 chronically HBV-infected European patients exposed to ≥ 1 NA, with detectable HBV-DNA and with an available HBsAg-sequence. The immune-associated escape mutations and the NA-induced immune-escape mutations sI195M, sI196S, and sE164D (resulting from drug-resistance mutation rtM204 V, rtM204I, and rtV173L) were retrieved from literature and examined. Mutations were defined as an aminoacid substitution with respect to a genotype A or D reference sequence. At least one immune-associated escape mutation was detected in 22.1% of patients with rising temporal-trend. By multivariable-analysis, genotype-D correlated with higher selection of ≥ 1 immune-associated escape mutation (OR[95%CI]:2.20[1.32-3.67], P = 0.002). In genotype-D, the presence of ≥ 1 immune-associated escape mutations was significantly higher in drug-exposed patients with drug-resistant strains than with wild-type virus (29.5% vs 20.3% P = 0.012). Result confirmed by analysing drug-naïve patients (29.5% vs 21.2%, P = 0.032). Strong correlation was observed between sP120T and rtM204I/V (P < 0.001), and their co-presence determined an increased HBV-DNA. At least one NA-induced immune-escape mutation occurred in 28.6% of patients, and their selection correlated with genotype-A (OR[95%CI]:2.03[1.32-3.10],P = 0.001). Finally

Insight into the molecular mechanism of yeast acetyl-coenzyme A carboxylase mutants F510I, N485G, I69E, E477R, and K73R resistant to soraphen A

NASA Astrophysics Data System (ADS)

Gao, Jian; Liang, Li; Chen, Qingqing; Zhang, Ling; Huang, Tonghui

2018-02-01

Acetyl-coenzyme A carboxylases (ACCs) is the first committed enzyme of fatty acid synthesis pathway. The inhibition of ACC is thought to be beneficial not only for diseases related to metabolism, such as type-2 diabetes, but also for infectious disease like bacterial infection disease. Soraphen A, a potent allosteric inhibitor of BC domain of yeast ACC, exhibit lower binding affinities to several yeast ACC mutants and the corresponding drug resistance mechanisms are still unknown. We report here a theoretical study of binding of soraphen A to wild type and yeast ACC mutants (including F510I, N485G, I69E, E477R, and K73R) via molecular dynamic simulation and molecular mechanics/generalized Born surface area free energy calculations methods. The calculated binding free energies of soraphen A to yeast ACC mutants are weaker than to wild type, which is highly consistent with the experimental results. The mutant F510I weakens the binding affinity of soraphen A to yeast ACC mainly by decreasing the van der Waals contributions, while the weaker binding affinities of Soraphen A to other yeast ACC mutants including N485G, I69E, E477R, and K73R are largely attributed to the decreased net electrostatic (ΔE ele + ΔG GB) interactions. Our simulation results could provide important insights for the development of more potent ACC inhibitors.

Hepatitis B surface antigen (HBsAg) and core antigen (HBcAg) combine CpG oligodeoxynucletides as a novel therapeutic vaccine for chronic hepatitis B infection.

PubMed

Li, Jianqiang; Ge, Jun; Ren, Sulin; Zhou, Tong; Sun, Ying; Sun, Honglin; Gu, Yue; Huang, Hongying; Xu, Zhenxing; Chen, Xiaoxiao; Xu, Xiaowei; Zhuang, Xiaoqian; Song, Cuiling; Jia, Fangmiao; Xu, Aiguo; Yin, Xiaojin; Du, Sean X

2015-08-20

Hepatitis B virus infection is a non-cytopathic hepatotropic virus which can lead to chronic liver disease and hepatocellular carcinoma. Traditional therapies fail to provide sustained control of viral replication and liver damage in most patients. As an alternative strategy, immunotherapeutic approaches have shown promising efficacy in the treatment of chronic hepatitis B patients. Here, we investigated the efficacy of a novel therapeutic vaccine formulation consisting of two HBV antigens, HBsAg and HBcAg, and CpG adjuvant. This vaccine formulation elicits forceful humoral responses directed against HBsAg/HBcAg, and promotes a Th1/Th2 balance response against HBsAg and a Th1-biased response against HBcAg in both C57BL/6 and HBV transgenic mice. Vigorous cellular immune response was also detected in HBV transgenic mice, for a significantly higher number of HBs/HBc-specific IFN-γ secreting CD4+ and CD8+ T cells was generated. Moreover, vaccinated mice elicited significantly intense in vivo CTL attack, reduced serum HBsAg level without causing liver damage in HBV transgenic mice. In summary, this study demonstrates a novel therapeutic vaccine with the potential to elicit vigorous humoral and cellular response, overcoming tolerance in HBV transgenic mice. Copyright © 2015 Elsevier Ltd. All rights reserved.

Expression of salt-induced 2-Cys peroxiredoxin from Oryza sativa increases stress tolerance and fermentation capacity in genetically engineered yeast Saccharomyces cerevisiae.

PubMed

Kim, Il-Sup; Kim, Young-Saeng; Yoon, Ho-Sung

2013-04-01

Peroxiredoxins (Prxs), also termed thioredoxin peroxidases (TPXs), are a family of thiol-specific antioxidant enzymes that are critically involved in cell defense and protect cells from oxidative damage. In this study, a putative chloroplastic 2-Cys thioredoxin peroxidase (OsTPX) was identified by proteome analysis from leaf tissue samples of rice (Oryza sativa) seedlings exposed to 0.1 M NaCl for 3 days. To investigate the relationship between the OsTPX gene and the stress response, OsTPX was cloned into the yeast expression vector p426GPD under the control of the glyceraldehyde-3-phosphate dehydrogenase (GPD1) promoter, and the construct was transformed into Saccharomyces cerevisiae cells. OsTPX expression was confirmed by semi-quantitative reverse transcription-polymerase chain reaction and western blot analyses. OsTPX contained two highly conserved cysteine residues (Cys114 and Cys236) and an active site region (FTFVCPT), and it is structurally very similar to human 2-Cys Prx. Heterologous OsTPX expression increased the ability of the transgenic yeast cells to adapt and recover from reactive oxygen species (ROS)-induced oxidative stresses, such as a reduction of cellular hydroperoxide levels in the presence of hydrogen peroxide and menadione, by improving redox homeostasis. OsTPX expression also conferred enhanced tolerance to tert-butylhydroperoxide, heat shock, and high ethanol concentrations. Furthermore, high OsTPX expression improved the fermentation capacity of the yeast during glucose-based batch fermentation at a high temperature (40 °C) and at the general cultivation temperature (30 °C). The alcohol yield in OsTPX-expressing transgenic yeast increased by approximately 29 % (0.14 g g(-1)) and 21 % (0.12 g g(-1)) during fermentation at 40 and 30 °C, respectively, compared to the wild-type yeast. Accordingly, OsTPX-expressing transgenic yeast showed prolonged cell survival during the environmental stresses produced during fermentation. These

Flavocoxid exerts a potent antiviral effect against hepatitis B virus.

PubMed

Pollicino, Teresa; Musolino, Cristina; Irrera, Natasha; Bitto, Alessandra; Lombardo, Daniele; Timmoneri, Martina; Minutoli, Letteria; Raimondo, Giovanni; Squadrito, Giovanni; Squadrito, Francesco; Altavilla, Domenica

2018-01-01

Flavocoxid is a proprietary blend of two flavonoids, baicalin and catechin, and recent evidence has shown that bioflavonoids may exert antiviral activities. The potential antiviral activity of Flavocoxid against hepatitis B virus (HBV) was evaluated. Additionally, it was investigated if Flavocoxid used in combination with Entecavir could potentiate its anti-HBV activity. Hepatoma cells replicating HBV were treated with Flavocoxid, or Entecavir alone or in combination for up to 5 days. Viral replicative intermediates, transcripts, and cccDNA levels were evaluated in HBV-replicating cells by real-time PCR, Southern and Northern blotting. Expression profiling was performed using TaqMan low-density arrays. Flavocoxid treatment induced a reduction of HBV replicative intermediates, the amount of transcripts, and HBsAg levels. Flavocoxid and Entecavir combination therapy further decreased the amount of HBV replicative intermediates, compared to Flavocoxid alone. Importantly, Flavocoxid alone or in combination with Entecavir also induced a reduction of cccDNA. Gene-expression analysis showed that Flavocoxid activates type I IFNs-signaling and dampens the HBV-induced inflammatory response. Flavocoxid inhibits HBV replication by targeting multiple steps of viral life cycle. These results indicate that the antiviral activity of Entecavir is potentiated by Flavocoxid, suggesting that this medical food might be considered as an adjuvant for anti-HBV therapy.

Improving industrial yeast strains: exploiting natural and artificial diversity.

PubMed

Steensels, Jan; Snoek, Tim; Meersman, Esther; Picca Nicolino, Martina; Voordeckers, Karin; Verstrepen, Kevin J

2014-09-01

Yeasts have been used for thousands of years to make fermented foods and beverages, such as beer, wine, sake, and bread. However, the choice for a particular yeast strain or species for a specific industrial application is often based on historical, rather than scientific grounds. Moreover, new biotechnological yeast applications, such as the production of second-generation biofuels, confront yeast with environments and challenges that differ from those encountered in traditional food fermentations. Together, this implies that there are interesting opportunities to isolate or generate yeast variants that perform better than the currently used strains. Here, we discuss the different strategies of strain selection and improvement available for both conventional and nonconventional yeasts. Exploiting the existing natural diversity and using techniques such as mutagenesis, protoplast fusion, breeding, genome shuffling and directed evolution to generate artificial diversity, or the use of genetic modification strategies to alter traits in a more targeted way, have led to the selection of superior industrial yeasts. Furthermore, recent technological advances allowed the development of high-throughput techniques, such as 'global transcription machinery engineering' (gTME), to induce genetic variation, providing a new source of yeast genetic diversity. © 2014 The Authors. FEMS Microbiology Reviews published by John Wiley & Sons Ltd on behalf of Federation of European Microbiological Societies.

Proteomic and metabolomic analysis of the carotenogenic yeast Xanthophyllomyces dendrorhous using different carbon sources.

PubMed

Martinez-Moya, Pilar; Niehaus, Karsten; Alcaíno, Jennifer; Baeza, Marcelo; Cifuentes, Víctor

2015-04-12

Astaxanthin is a potent antioxidant with increasing biotechnological interest. In Xanthophyllomyces dendrorhous, a natural source of this pigment, carotenogenesis is a complex process regulated through several mechanisms, including the carbon source. X. dendrorhous produces more astaxanthin when grown on a non-fermentable carbon source, while decreased astaxanthin production is observed in the presence of high glucose concentrations. In the present study, we used a comparative proteomic and metabolomic analysis to characterize the yeast response when cultured in minimal medium supplemented with glucose (fermentable) or succinate (non-fermentable). A total of 329 proteins were identified from the proteomic profiles, and most of these proteins were associated with carotenogenesis, lipid and carbohydrate metabolism, and redox and stress responses. The metabolite profiles revealed 92 metabolites primarily associated with glycolysis, the tricarboxylic acid cycle, amino acids, organic acids, sugars and phosphates. We determined the abundance of proteins and metabolites of the central pathways of yeast metabolism and examined the influence of these molecules on carotenogenesis. Similar to previous proteomic-stress response studies, we observed modulation of abundance from several redox, stress response, carbohydrate and lipid enzymes. Additionally, the accumulation of trehalose, absence of key ROS response enzymes, an increased abundance of the metabolites of the pentose phosphate pathway and tricarboxylic acid cycle suggested an association between the accumulation of astaxanthin and oxidative stress in the yeast. Moreover, we observed the increased abundance of late carotenogenesis enzymes during astaxanthin accumulation under succinate growth conditions. The use of succinate as a carbon source in X. dendrorhous cultures increases the availability of acetyl-CoA for the astaxanthin production compared with glucose, likely reflecting the positive regulation of metabolic

Sodium orthovanadate (vanadate), a potent mitigator of radiation-induced damage to the hematopoietic system in mice

PubMed Central

Wang, Bing; Tanaka, Kaoru; Morita, Akinori; Ninomiya, Yasuharu; Maruyama, Kouichi; Fujita, Kazuko; Hosoi, Yoshio; Nenoi, Mitsuru

2013-01-01

Previous in vitro and in vivo studies have shown that sodium orthovanadate (vanadate), an inorganic vanadium compound, could effectively suppress radiation-induced p53-mediated apoptosis via both transcription-dependent and transcription-independent pathways. As a potent radiation protector administered at a dose of 20 mg/kg body weight (20 mg/kg) prior to total body irradiation (TBI) by intra-peritoneal (ip) injection, it completely protected mice from hematopoietic syndrome and partially from gastrointestinal syndrome. In the present study, radiation mitigation effects from vanadate were investigated by ip injection of vanadate after TBI in mice. Results showed that a single administration of vanadate at a dose of 20 mg/kg markedly improved the 30-day survival rate and the peripheral blood hemogram, relieved bone marrow aplasia and decreased occurrence of the bone marrow micronucleated erythrocytes in the surviving animals. The dose reduction factor was 1.2 when a single dose of 20 mg/kg was administered 15 min after TBI in mice using the 30-day survival test as the endpoint. Results also showed that either doubling the vanadate dose (40 mg/kg) in a single administration or continuing the vanadate treatment (after a single administration at 20 mg/kg) from the following day at a dose of 5 mg/kg per day for 4 consecutive days further significantly improved the efficacy for rescuing bone marrow failure in the 30-day survival test. Taken together, these findings indicate that vanadate would be a potent mitigator suppressing the acute lethality (hematopoietic syndrome) and minimizing the detrimental effects (anhematopoiesis and delayed genotoxic effects) induced by TBI in mice. PMID:23349341

Kex1 protease is involved in yeast cell death induced by defective N-glycosylation, acetic acid, and chronological aging.

PubMed

Hauptmann, Peter; Lehle, Ludwig

2008-07-04

N-glycosylation in the endoplasmic reticulum is an essential protein modification and highly conserved in evolution from yeast to humans. The key step of this pathway is the transfer of the lipid-linked core oligosaccharide to the nascent polypeptide chain, catalyzed by the oligosaccharyltransferase complex. Temperature-sensitive oligosaccharyltransferase mutants of Saccharomyces cerevisiae at the restrictive temperature, such as wbp1-1, as well as wild-type cells in the presence of the N-glycosylation inhibitor tunicamycin display typical apoptotic phenotypes like nuclear condensation, DNA fragmentation, phosphatidylserine translocation, caspase-like activity, and reactive oxygen species accumulation. Since deletion of the yeast metacaspase YCA1 did not abrogate this death pathway, we postulated a different proteolytic process to be responsible. Here, we show that Kex1 protease is involved in the programmed cell death caused by defective N-glycosylation. Its disruption decreases caspase-like activity, production of reactive oxygen species, and fragmentation of mitochondria and, conversely, improves growth and survival of cells. Moreover, we demonstrate that Kex1 contributes also to the active cell death program induced by acetic acid stress or during chronological aging, suggesting that Kex1 plays a more general role in cellular suicide of yeast.

Epidemiology and antifungal susceptibilities of yeasts causing vulvovaginitis in a teaching hospital.

PubMed

Gamarra, Soledad; Morano, Susana; Dudiuk, Catiana; Mancilla, Estefanía; Nardin, María Elena; de Los Angeles Méndez, Emilce; Garcia-Effron, Guillermo

2014-10-01

Vulvovaginal candidiasis is one of the most common mycosis. However, the information about antifungal susceptibilities of the yeasts causing this infection is scant. We studied 121 yeasts isolated from 118 patients with vulvovaginal candidiasis. The isolates were identified by phenotypic and molecular methods, including four phenotypic methods described to differentiate Candida albicans from C. dubliniensis. Antifungal susceptibility testing was performed according to CLSI documents M27A3 and M27S4 using the drugs available as treatment option in the hospital. Diabetes, any antibacterial and amoxicillin treatment were statistically linked with vulvovaginal candidiasis, while oral contraceptives were not considered a risk factor. Previous azole-based over-the-counter antifungal treatment was statistically associated with non-C.albicans yeasts infections. The most common isolated yeast species was C. albicans (85.2 %) followed by C. glabrata (5 %), Saccharomyces cerevisiae (3.3 %), and C. dubliniensis (2.5 %). Fluconazole- and itraconazole-reduced susceptibility was observed in ten and in only one C. albicans strains, respectively. All the C. glabrata isolates showed low fluconazole MICs. Clotrimazole showed excellent potency against all but seven isolates (three C. glabrata, two S. cerevisiae, one C. albicans and one Picchia anomala). Any of the strains showed nystatin reduced susceptibility. On the other hand, terbinafine was the less potent drug. Antifungal resistance is still a rare phenomenon supporting the use of azole antifungals as empirical treatment of vulvovaginal candidiasis.

Calcitonin-gene related peptide is a potent inducer of oedema in rat orofacial tissue.

PubMed

Queiroz, Bárbara F G de; Almeida, Marcella P A de; Bakhle, Y S; Francischi, Janetti N

2018-04-01

This study aimed to assess the potential of calcitonin-gene related peptide (CGRP), a neuropeptide released from sensory nerves, to induce oedema in orofacial tissue. Wistar rats (150-200 g) anesthetized with isoflurane were injected intraorally with CGRP (100 μl; 8-33 pmol) in the right side of the mouth. The contralateral side was injected with the same volume of physiological saline. Increased cheek thickness (in mm), as a measure of oedema formation, was assayed bilaterally with a digital caliper before (T = 0) and up to 24 h following injection of CGRP. Pretreatment with antagonists (CGRP 8-37, 10 nmol; pizotifen, 2 mg/kg) was given by intra-oral or subcutaneous injection, 10 or 30 min, respectively, before the inflammatory stimulus. CGRP and CGRP 8-37 were also injected into the rat hind paw to induce oedema. Data are presented as the mean (±SEM) difference in thickness between the right and the left sides at each time. Following intra-oral injection, CGRP induced a rapidly developing (5-15 min) and long-lasting (6 h), dose-dependent oedema in the rat cheek, blocked by pre-treatment with CGRP 8-37 or pizotifen. CGRP induced a smaller oedematogenic effect in the rat hind paw also blocked by the CGRP antagonist. CGRP (16 pmol) potentiated the oedema induced by co-injected substance P (3.7 nmol) and contributed to the oedema following intraoral injection of carrageenan (100 μg). Injection of CGRP 8-37 alone induced an early but short-lasting oedema. Local injection of CGRP potently induced oedema in the orofacial tissue of rats which was blocked by a CGRP receptor antagonist. The overall inhibition of carrageenan-induced oedema by CGRP 8-37 suggests that endogenous CGRP contributes to an oedematogenic response in orofacial tissues. Copyright © 2018 Elsevier Ltd. All rights reserved.

An NQO1 substrate with potent antitumor activity that selectively kills by PARP1-induced programmed necrosis.

PubMed

Huang, Xiumei; Dong, Ying; Bey, Erik A; Kilgore, Jessica A; Bair, Joseph S; Li, Long-Shan; Patel, Malina; Parkinson, Elizabeth I; Wang, Yiguang; Williams, Noelle S; Gao, Jinming; Hergenrother, Paul J; Boothman, David A

2012-06-15

Agents, such as β-lapachone, that target the redox enzyme, NAD(P)H:quinone oxidoreductase 1 (NQO1), to induce programmed necrosis in solid tumors have shown great promise, but more potent tumor-selective compounds are needed. Here, we report that deoxynyboquinone kills a wide spectrum of cancer cells in an NQO1-dependent manner with greater potency than β-lapachone. Deoxynyboquinone lethality relies on NQO1-dependent futile redox cycling that consumes oxygen and generates extensive reactive oxygen species (ROS). Elevated ROS levels cause extensive DNA lesions, PARP1 hyperactivation, and severe NAD+ /ATP depletion that stimulate Ca2+ -dependent programmed necrosis, unique to this new class of NQO1 "bioactivated" drugs. Short-term exposure of NQO1+ cells to deoxynyboquinone was sufficient to trigger cell death, although genetically matched NQO1- cells were unaffected. Moreover, siRNA-mediated NQO1 or PARP1 knockdown spared NQO1+ cells from short-term lethality. Pretreatment of cells with BAPTA-AM (a cytosolic Ca2+ chelator) or catalase (enzymatic H2O2 scavenger) was sufficient to rescue deoxynyboquinone-induced lethality, as noted with β-lapachone. Investigations in vivo showed equivalent antitumor efficacy of deoxynyboquinone to β-lapachone, but at a 6-fold greater potency. PARP1 hyperactivation and dramatic ATP loss were noted in the tumor, but not in the associated normal lung tissue. Our findings offer preclinical proof-of-concept for deoxynyboquinone as a potent chemotherapeutic agent for treatment of a wide spectrum of therapeutically challenging solid tumors, such as pancreatic and lung cancers.

Prequels to Synthetic Biology: From Candidate Gene Identification and Validation to Enzyme Subcellular Localization in Plant and Yeast Cells.

PubMed

Foureau, E; Carqueijeiro, I; Dugé de Bernonville, T; Melin, C; Lafontaine, F; Besseau, S; Lanoue, A; Papon, N; Oudin, A; Glévarec, G; Clastre, M; St-Pierre, B; Giglioli-Guivarc'h, N; Courdavault, V

2016-01-01

Natural compounds extracted from microorganisms or plants constitute an inexhaustible source of valuable molecules whose supply can be potentially challenged by limitations in biological sourcing. The recent progress in synthetic biology combined to the increasing access to extensive transcriptomics and genomics data now provide new alternatives to produce these molecules by transferring their whole biosynthetic pathway in heterologous production platforms such as yeasts or bacteria. While the generation of high titer producing strains remains per se an arduous field of investigation, elucidation of the biosynthetic pathways as well as characterization of their complex subcellular organization are essential prequels to the efficient development of such bioengineering approaches. Using examples from plants and yeasts as a framework, we describe potent methods to rationalize the study of partially characterized pathways, including the basics of computational applications to identify candidate genes in transcriptomics data and the validation of their function by an improved procedure of virus-induced gene silencing mediated by direct DNA transfer to get around possible resistance to Agrobacterium-delivery of viral vectors. To identify potential alterations of biosynthetic fluxes resulting from enzyme mislocalizations in reconstituted pathways, we also detail protocols aiming at characterizing subcellular localizations of protein in plant cells by expression of fluorescent protein fusions through biolistic-mediated transient transformation, and localization of transferred enzymes in yeast using similar fluorescence procedures. Albeit initially developed for the Madagascar periwinkle, these methods may be applied to other plant species or organisms in order to establish synthetic biology platform. © 2016 Elsevier Inc. All rights reserved.

Poly (I:C) enhances the anti-tumor activity of canine parvovirus NS1 protein by inducing a potent anti-tumor immune response.

PubMed

Gupta, Shishir Kumar; Yadav, Pavan Kumar; Tiwari, A K; Gandham, Ravi Kumar; Sahoo, A P

2016-09-01

The canine parvovirus NS1 (CPV2.NS1) protein selectively induces apoptosis in the malignant cells. However, for an effective in vivo tumor treatment strategy, an oncolytic agent also needs to induce a potent anti-tumor immune response. In the present study, we used poly (I:C), a TLR3 ligand, as an adjuvant along with CPV2.NS1 to find out if the combination can enhance the oncolytic activity by inducing a potent anti-tumor immune response. The 4T1 mammary carcinoma cells were used to induce mammary tumor in Balb/c mice. The results suggested that poly (I:C), when given along with CPV2.NS1, not only significantly reduced the tumor growth but also augmented the immune response against tumor antigen(s) as indicated by the increase in blood CD4+ and CD8+ counts and infiltration of immune cells in the tumor tissue. Further, blood serum analysis of the cytokines revealed that Th1 cytokines (IFN-γ and IL-2) were significantly upregulated in the treatment group indicating activation of cell-mediated immune response. The present study reports the efficacy of CPV2.NS1 along with poly (I:C) not only in inhibiting the mammary tumor growth but also in generating an active anti-tumor immune response without any visible toxicity. The results of our study may help in developing CPV2.NS1 and poly (I: C) combination as a cancer therapeutic regime to treat various malignancies.

The timing of UV mutagenesis in yeast: a pedigree analysis of induced recessive mutation.

PubMed

James, A P; Kilbey, B J

1977-10-01

The mechanism of UV-induced mutation in eukaryotes was studied in individual yeast cells by a procedure that combined pedigree analysis and tetrad analysis. The technique involved the induction of recessive lethals and semilethals in G1 diploid cells. Induced frequencies were 25 and 61 percent at survival levels of 90 and 77 percent, respectively. No evidence of gross chromosome aberrations was detected. Recessive mutations that affect only one strand or that affect both strands of the DNA molecule are induced much at random among a population of cells, and both types can occur within the same cell. However, the data confirm that two-strand mutations are in the majority after a low level of irradiation. The simplest explanation involves a mechanism whereby most mutations are fixed in both strands prior to the first round of post-irradiation DNA replication. The recessive mutational consequences of irradiation are exhausted at the conclusion of the first post-irradiation cell division, although dominant-lethal sectoring continues at a high level through the second post-irradiation division. It is concluded that pyrimidine dimers that persist to the second round of DNA replication are rare or ineffective.

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.

Potent Antiarthritic Properties of Phloretin in Murine Collagen-Induced Arthritis.

PubMed

Wang, Shun-Ping; Lin, Shih-Chao; Li, Shiming; Chao, Ya-Hsuan; Hwang, Guang-Yuh; Lin, Chi-Chen

2016-01-01

In the exploration of potential therapeutic agents for rheumatoid arthritis (RA), DBA/1J mice are used as the RA model of collagen-induced arthritis (CIA). Phloretin, a flavonoid compound extracted from Prunus mandshurica , has been found to exhibit anti-inflammatory activity, making it a potential candidate for treatment of RA. The objective of this study was to evaluate the therapeutic effects of phloretin on CIA mice. CIA mice were dosed daily with phloretin at either 50 or 100 mg/kg among two treatment groups. CIA treated mice showed mitigation of clinical symptoms of RA in addition to reduced inflammation of hind-limbs compared to mice who did not receive phloretin. Histological analysis showed that phloretin suppressed the severity of RA and effectively mitigated joint inflammation and cartilage- and bone-destruction via reducing proinflammatory cytokine productions (TNF- α , IL-6, IL-1 β , and IL-17). This was at least partially mediated by causing inadequate splenocyte activation and proliferation. Moreover, phloretin-treated CIA mice showed decreased oxidative stress and diminished levels of malondialdehyde (MDA) and hydrogen peroxide (H 2 O 2 ) in paw tissues as well as reduced productivity of anti-collagen antibodies in serum. We have concluded that phloretin could be a potent and effective antiarthritis agent, demonstrating anti-inflammatory, antioxidative, and immunomodulatory effects in CIA mice.

Synthetic genome engineering forging new frontiers for wine yeast.

PubMed

Pretorius, Isak S

2017-02-01

Over the past 15 years, the seismic shifts caused by the convergence of biomolecular, chemical, physical, mathematical, and computational sciences alongside cutting-edge developments in information technology and engineering have erupted into a new field of scientific endeavor dubbed Synthetic Biology. Recent rapid advances in high-throughput DNA sequencing and DNA synthesis techniques are enabling the design and construction of new biological parts (genes), devices (gene networks) and modules (biosynthetic pathways), and the redesign of biological systems (cells and organisms) for useful purposes. In 2014, the budding yeast Saccharomyces cerevisiae became the first eukaryotic cell to be equipped with a fully functional synthetic chromosome. This was achieved following the synthesis of the first viral (poliovirus in 2002 and bacteriophage Phi-X174 in 2003) and bacterial (Mycoplasma genitalium in 2008 and Mycoplasma mycoides in 2010) genomes, and less than two decades after revealing the full genome sequence of a laboratory (S288c in 1996) and wine (AWRI1631 in 2008) yeast strain. A large international project - the Synthetic Yeast Genome (Sc2.0) Project - is now underway to synthesize all 16 chromosomes (∼12 Mb carrying ∼6000 genes) of the sequenced S288c laboratory strain by 2018. If successful, S. cerevisiae will become the first eukaryote to cross the horizon of in silico design of complex cells through de novo synthesis, reshuffling, and editing of genomes. In the meantime, yeasts are being used as cell factories for the semi-synthetic production of high-value compounds, such as the potent antimalarial artemisinin, and food ingredients, such as resveratrol, vanillin, stevia, nootkatone, and saffron. As a continuum of previously genetically engineered industrially important yeast strains, precision genome engineering is bound to also impact the study and development of wine yeast strains supercharged with synthetic DNA. The first taste of what the future

Discovery and structure-activity relationships of (2-(arylthio)benzylideneamino)guanidines as a novel series of potent apoptosis inducers.

PubMed

Zhang, Han-Zhong; Crogan-Grundy, Candace; May, Chris; Drewe, John; Tseng, Ben; Cai, Sui Xiong

2009-04-01

1-(2-(2,5-Dimethoxyphenylthio)benzylidene)semicarbazide (2a) was discovered as a potent apoptosis inducer through our cell based HTS assay. SAR study led to the discovery of a more aqueous soluble analog (2-(2,5-dimethoxyphenylthio)-6-methoxybenzylideneamino)guanidine (5e) with EC(50) value of 60 nM in the caspase activation assay and GI(50) value of 62 nM in the growth inhibition assay in T47D cells. Compound 5e was found to be an inhibitor of tubulin polymerization and efficacious in a MX-1 breast tumor model.

Design, synthesis and SARs of novel salicylanilides as potent inhibitors of RANKL-induced osteoclastogenesis and bone resorption.

PubMed

Chen, Chun-Liang; Lee, Chia-Chung; Liu, Fei-Lan; Chen, Tsung-Chih; Ahmed Ali, Ahmed Atef; Chang, Deh-Ming; Huang, Hsu-Shan

2016-07-19

Inhibiting osteoclastogenesis is a promising therapeutic target for treating osteoclast-related diseases. Herein, we synthesized a series of modified salicylanilides and their corresponding 3-phenyl-2H-benzo[e][1,3]oxazine-2,4(3H)-dione and 10-phenyldibenzo[b,f][1,4]oxazepin-11(10H)-one derivatives, and investigated the effects of such compounds on RANKL-induced osteoclast formation. Among them, a salicylanilide derivative (A04) and its 3-phenyl-2H-benzo[e][1,3]oxazine-2,4(3H)-dione derivative (B04) markedly suppressed RANKL-induced osteoclast differentiation and showed no significant cytotoxic effects at doses higher than that required to inhibit osteoclast formation. Both compounds reduced osteoclast formation and bone resorptive activity of osteoclasts in a dose-dependent manner. Further, the anti-osteoclastogenic effects of A04 and B04 may operate through reducing the RANKL-induced nuclear translocation of NFATc1. Accordingly, we present the potent anti-osteoclastogenic compounds A04 and B04 as promising candidates for further optimization as anti-resorptive agents. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Engineering of synthetic, stress-responsive yeast promoters

PubMed Central

Rajkumar, Arun S.; Liu, Guodong; Bergenholm, David; Arsovska, Dushica; Kristensen, Mette; Nielsen, Jens; Jensen, Michael K.; Keasling, Jay D.

2016-01-01

Advances in synthetic biology and our understanding of the rules of promoter architecture have led to the development of diverse synthetic constitutive and inducible promoters in eukaryotes and prokaryotes. However, the design of promoters inducible by specific endogenous or environmental conditions is still rarely undertaken. In this study, we engineered and characterized a set of strong, synthetic promoters for budding yeast Saccharomyces cerevisiae that are inducible under acidic conditions (pH ≤ 3). Using available expression and transcription factor binding data, literature on transcriptional regulation, and known rules of promoter architecture we improved the low-pH performance of the YGP1 promoter by modifying transcription factor binding sites in its upstream activation sequence. The engineering strategy outlined for the YGP1 promoter was subsequently applied to create a response to low pH in the unrelated CCW14 promoter. We applied our best promoter variants to low-pH fermentations, enabling ten-fold increased production of lactic acid compared to titres obtained with the commonly used, native TEF1 promoter. Our findings outline and validate a general strategy to iteratively design and engineer synthetic yeast promoters inducible to environmental conditions or stresses of interest. PMID:27325743

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.

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

An exceptionally potent inducer of cytoprotective enzymes: elucidation of the structural features that determine inducer potency and reactivity with Keap1.

PubMed

Dinkova-Kostova, Albena T; Talalay, Paul; Sharkey, John; Zhang, Ying; Holtzclaw, W David; Wang, Xiu Jun; David, Emilie; Schiavoni, Katherine H; Finlayson, Stewart; Mierke, Dale F; Honda, Tadashi

2010-10-29

The Keap1/Nrf2/ARE pathway controls a network of cytoprotective genes that defend against the damaging effects of oxidative and electrophilic stress, and inflammation. Induction of this pathway is a highly effective strategy in combating the risk of cancer and chronic degenerative diseases, including atherosclerosis and neurodegeneration. An acetylenic tricyclic bis(cyano enone) bearing two highly electrophilic Michael acceptors is an extremely potent inducer in cells and in vivo. We demonstrate spectroscopically that both cyano enone functions of the tricyclic molecule react with cysteine residues of Keap1 and activate transcription of cytoprotective genes. Novel monocyclic cyano enones, representing fragments of rings A and C of the tricyclic compound, reveal that the contribution to inducer potency of the ring C Michael acceptor is much greater than that of ring A, and that potency is further enhanced by spatial proximity of an acetylenic function. Critically, the simultaneous presence of two cyano enone functions in rings A and C within a rigid three-ring system results in exceptionally high inducer potency. Detailed understanding of the structural elements that contribute to the reactivity with the protein sensor Keap1 and to high potency of induction is essential for the development of specific and selective lead compounds as clinically relevant chemoprotective agents.

Ultra-wide band electromagnetic radiation does not affect UV-induced recombination and mutagenesis in yeast.

PubMed

Pakhomova, O N; Belt, M L; Mathur, S P; Lee, J C; Akyel, Y

1998-01-01

Cell samples of the yeast Saccharomyces cerevisiae were exposed to 100 J/m2 of 254 nm ultraviolet (UV) radiation followed by a 30 min treatment with ultra-wide band (UWB) electromagnetic pulses. The UWB pulses (101-104 kV/m, 1.0 ns width, 165 ps rise time) were applied at the repetition rates of 0 Hz (sham), 16 Hz, or 600 Hz. The effect of exposures was evaluated from the colony-forming ability of the cells on complete and selective media and the number of aberrant colonies. The experiments established no effect of UWB exposure on the UV-induced reciprocal and non-reciprocal recombination, mutagenesis, or cell survival.

Ergothioneine production using Methylobacterium species, yeast, and fungi.

PubMed

Fujitani, Yoshiko; Alamgir, Kabir Md; Tani, Akio

2018-06-14

Ergothioneine (EGT) is a sulfur-containing, anti-oxidative amino acid derived from histidine. EGT is synthesized in bacteria and fungi but not in animals and plants, and is now recognized as important for human health. Its cost-effective fermentative production has not been elucidated due to the lack of information for productive microorganisms. In this study, we doubled the gene copy for EGT synthesis and deleted the histidine ammonia-lyase gene in a potent EGT-producing methylotrophic bacterium Methylobacterium aquaticum strain 22A, and optimized its culture conditions, resulting in increased EGT production of 7.0 mg EGT/g dry cell weight and 100 μg EGT/5 mL/7 days. In addition, through screening we found EGT-producing eukaryotic strains of Aureobasidium pullulans and Rhodotorula mucilaginosa, which can produce 1.0 and 3.2 mg EGT/g dry cell weight, 70 and 120 μg EGT/5 mL/7 days, respectively. This study proposes practical uses of potent EGT-producing recombinant Methylobacterium species and non-recombinant yeast and fungal strains. Copyright © 2018 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

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.

Yeast Infection (Vaginal)

MedlinePlus

Yeast infection (vaginal) Overview A vaginal yeast infection is a fungal infection that causes irritation, discharge and intense itchiness ... symptoms Causes The fungus candida causes a vaginal yeast infection. Your vagina naturally contains a balanced mix of yeast, including ...

Breast milk immune complexes are potent inducers of oral tolerance in neonates and prevent asthma development.

PubMed

Mosconi, E; Rekima, A; Seitz-Polski, B; Kanda, A; Fleury, S; Tissandie, E; Monteiro, R; Dombrowicz, D D; Julia, V; Glaichenhaus, N; Verhasselt, V

2010-09-01

Allergic asthma is a chronic lung disease resulting from an inappropriate T helper (Th)-2 response to environmental antigens. Early tolerance induction is an attractive approach for primary prevention of asthma. Here, we found that breastfeeding by antigen-sensitized mothers exposed to antigen aerosols during lactation induced a robust and long-lasting antigen-specific protection from asthma. Protection was more profound and persistent than the one induced by antigen-exposed non-sensitized mothers. Milk from antigen-exposed sensitized mothers contained antigen-immunoglobulin (Ig) G immune complexes that were transferred to the newborn through the neonatal Fc receptor resulting in the induction of antigen-specific FoxP3(+) CD25(+) regulatory T cells. The induction of oral tolerance by milk immune complexes did not require the presence of transforming growth factor-beta in milk in contrast to tolerance induced by milk-borne free antigen. Furthermore, neither the presence of IgA in milk nor the expression of the inhibitory FcgammaRIIb in the newborn was required for tolerance induction. This study provides new insights on the mechanisms of tolerance induction in neonates and highlights that IgG immune complexes found in breast milk are potent inducers of oral tolerance. These observations may pave the way for the identification of key factors for primary prevention of immune-mediated diseases such as asthma.

iAID: an improved auxin-inducible degron system for the construction of a 'tight' conditional mutant in the budding yeast Saccharomyces cerevisiae.

PubMed

Tanaka, Seiji; Miyazawa-Onami, Mayumi; Iida, Tetsushi; Araki, Hiroyuki

2015-08-01

Isolation of a 'tight' conditional mutant of a gene of interest is an effective way of studying the functions of essential genes. Strategies that use ubiquitin-mediated protein degradation to eliminate the product of a gene of interest, such as heat-inducible degron (td) and auxin-inducible degron (AID), are powerful methods for constructing conditional mutants. However, these methods do not work with some genes. Here, we describe an improved AID system (iAID) for isolating tight conditional mutants in the budding yeast Saccharomyces cerevisiae. In this method, transcriptional repression by the 'Tet-OFF' promoter is combined with proteolytic elimination of the target protein by the AID system. To provide examples, we describe the construction of tight mutants of the replication factors Dpb11 and Mcm10, dpb11-iAID, and mcm10-iAID. Because Dpb11 and Mcm10 are required for the initiation of DNA replication, their tight mutants are unable to enter S phase. This is the case for dpb11-iAID and mcm10-iAID cells after the addition of tetracycline and auxin. Both the 'Tet-OFF' promoter and the AID system have been shown to work in model eukaryotes other than budding yeast. Therefore, the iAID system is not only useful in budding yeast, but also can be applied to other model systems to isolate tight conditional mutants. Copyright © 2015 John Wiley & Sons, Ltd.

Human immunodeficiency virus (HIV) seropositivity and hepatitis B surface antigenemia (HBSAG) among blood donors in Benin city, Edo state, Nigeria.

PubMed

Umolu, Patience Idia; Okoror, Lawrence Ehis; Orhue, Philip

2005-03-01

Human Immunodeficiency Virus and Hepatitis B virus are blood borne pathogens that can be transmitted through blood transfusion and could pose a huge problem in areas where mechanisms of ensuring blood safety are suspect. This study became necessary in a population where most of the blood for transfusion is from commercial blood donors. A total of 130 donors comprising 120 commercial donors and 10 voluntary donors were tested for antibodies to human immunodeficiency virus and hepatitis B surface antigen in Benin city using Immunocomb HIV - 1 and 2 Biospot kit and Quimica Clinica Aplicada direct latex agglutination method respectively. Thirteen (10%) samples were HIV seropositive and 7(5.8%) were HBsAg positive. The age bracket 18 - 25years had the highest numbers of donors and also had the highest number of HBsAg positive cases (7.8%) while the age group 29 - 38years had highest number of HIV seropositive cases. High prevalence of HIV antibodies and Hepatitis B surface antigen was found among commercial blood donors. Appropriate and compulsory screening of blood donors using sensitive methods, must be ensured to prevent post transfusion hepatitis and HIV.

Changes in the relative population size of selected ruminal bacteria following an induced episode of acidosis in beef heifers receiving viable and non-viable active dried yeast.

PubMed

Mohammed, R; Vyas, D; Yang, W Z; Beauchemin, K A

2017-06-01

To characterize the changes in the relative population size (RPS) of select ruminal bacteria and rumen fermentation variables in beef heifers supplemented with a strain of Saccharomyces cerevisiae as viable active dried (ADY) or killed dried (KDY) yeast following an induced episode of ruminal acidosis. Six ruminally cannulated beef heifers fed a diet consisting of 50% forage and 50% grain (dry matter basis) were used in a replicated 3 × 3 Latin square design with three 28-day periods. Treatments were: (i) control (CTRL; no yeast); (ii) ADY (4 g day -1 providing 10 10  CFU per g; AB Vista, UK); and (iii) KDY (4 g day -1 autoclaved ADY). The acidosis challenge was induced on day 22 and rumen samples were collected on day 15 (baseline; BASE), day 22 (challenge day; CHAL), and on day 29 (168th hour post acid challenge or recovery, REC) of each period. Over the study, duration of pH <5·8 (indicative of subacute ruminal acidosis) was less for ADY and KDY than CTRL, with ADY less than KDY. No treatment effects were observed on relative abundance of ruminal bacteria, but the day effect was significant. The RPS of lactate producers and utilizers was greater while RPS of fibrolytic bacteria was lower during CHAL than BASE and REC. Yeast supplementation, irrespective of its viability, showed beneficial effects on ruminal pH variables in animals more susceptible to acidosis. Rumen microbial population was altered with the induction of severe acidosis. Most of the changes reverted back to baseline values during the recovery phase. Yeast supplementation reduced subacute rumen acidosis in the most susceptible cattle, but failed to attenuate severe acidosis induced by a grain challenge. The study provided valuable insight into the mechanism by which acidosis affects cattle performance. Individual animal variation in ruminal fermentation partly explained the variability in response to yeast supplementation in the study. © 2017 Her Majesty the Queen in Right of Canada

Mutations in the S gene and in the overlapping reverse transcriptase region in chronic hepatitis B Chinese patients with coexistence of HBsAg and anti-HBs.

PubMed

Ding, Feng; Miao, Xi-Li; Li, Yan-Xia; Dai, Jin-Fen; Yu, Hong-Gang

2016-01-01

The mechanism underlying the coexistence of hepatitis B surface antigen and antibodies to HBsAg in chronic hepatitis B patients remains unknown. This research aimed to determine the clinical and virological features of the rare pattern. A total of 32 chronic hepatitis B patients infected by HBV genotype C were included: 15 carrying both HBsAg and anti-HBs (group I) and 17 solely positive for HBsAg (group II). S gene and reverse transcriptase region sequences were amplified, sequenced and compared with the reference sequences. The amino acid variability within major hydrophilic region, especially the "a" determinant region, and within reverse transcriptase for regions overlapping the major hydrophilic region in group I is significantly higher than those in group II. Mutation sI126S/T within the "a" determinant was the most frequent change, and only patients from group I had the sQ129R, sG130N, sF134I, sG145R amino acid changes, which are known to alter immunogenicity. In chronic patients, the concurrent HBsAg/anti-HBs serological profile is associated with an increased aa variability in several key areas of HBV genome. Additional research on these genetic mutants are needed to clarify their biological significance for viral persistence. Copyright © 2015 Elsevier Editora Ltda. All rights reserved.

Characterization of a potent platelet aggregation inducer from Cerastes cerastes (Egyptian sand viper) venom.

PubMed

Basheer, A R; el-Asmar, M F; Soslau, G

1995-07-03

A potent, proteinaceous inducer of platelet aggregation designated as IVa, has been purified to homogeneity from Cerastes cerastes venom by molecular sieve and ion exchange chromatography. It is composed of 2 subunits with total M(r) of 62,000 as shown by native gel chromatography and chemical cross-linking with disuccinimidyl suberate. It is not clear at the present time whether both subunits are identical gene products, however, both have identical N-terminal sequences for the first 15 amino acids. The protein has a pI above 9.6. IVa (0.1 micrograms/ml) could aggregate platelets up to 80% and was inhibited by p-APMSF, leupeptin, iodoacetamide, protein kinase C inhibitor, phosphatase inhibitor, ATP and PGE1, while it was insensitive to acetylsalicylic acid, ADP scavenger system, protein kinase A inhibitor and hirudin. Protein IVa is a serine proteinase with thrombin-like activity as it hydrolysed thrombin chromogenic substrate CBS 34.47, its aggregatory activity was partially inhibited by monoclonal antibodies against GPIb and the thrombin receptor, as was the thrombin, and its ability to induce intracellular Ca2+ release was blocked by pretreating platelets with thrombin. Unlike thrombin, the IVa protein showed very weak coagulant activity as indicated by plasma recalcification time and fibrinogen clotting time although it could hydrolyse fibrinogen alpha-chains.

Distinct Domestication Trajectories in Top-Fermenting Beer Yeasts and Wine Yeasts.

PubMed

Gonçalves, Margarida; Pontes, Ana; Almeida, Pedro; Barbosa, Raquel; Serra, Marta; Libkind, Diego; Hutzler, Mathias; Gonçalves, Paula; Sampaio, José Paulo

2016-10-24

Beer is one of the oldest alcoholic beverages and is produced by the fermentation of sugars derived from starches present in cereal grains. Contrary to lager beers, made by bottom-fermenting strains of Saccharomyces pastorianus, a hybrid yeast, ale beers are closer to the ancient beer type and are fermented by S. cerevisiae, a top-fermenting yeast. Here, we use population genomics to investigate (1) the closest relatives of top-fermenting beer yeasts; (2) whether top-fermenting yeasts represent an independent domestication event separate from those already described; (3) whether single or multiple beer yeast domestication events can be inferred; and (4) whether top-fermenting yeasts represent non-recombinant or recombinant lineages. Our results revealed that top-fermenting beer yeasts are polyphyletic, with a main clade composed of at least three subgroups, dominantly represented by the German, British, and wheat beer strains. Other beer strains were phylogenetically close to sake, wine, or bread yeasts. We detected genetic signatures of beer yeast domestication by investigating genes previously linked to brewing and using genome-wide scans. We propose that the emergence of the main clade of beer yeasts is related with a domestication event distinct from the previously known cases of wine and sake yeast domestication. The nucleotide diversity of the main beer clade more than doubled that of wine yeasts, which might be a consequence of fundamental differences in the modes of beer and wine yeast domestication. The higher diversity of beer strains could be due to the more intense and different selection regimes associated to brewing. Copyright © 2016 Elsevier Ltd. All rights reserved.

Effect of a new potent CCK antagonist, lorglumide, on caerulein- and bombesin-induced pancreatic secretion and growth in the rat.

PubMed

Scarpignato, C; Varga, G; Dobronyi, I; Papp, M

1989-03-01

1. The effect of lorglumide, a new potent cholecystokinin (CCK) antagonist, on pancreatic secretion and growth induced by caerulein and bombesin was studied in the rat. 2. Pancreatic exocrine secretion was studied both in vitro (isolated and perfused pancreatic segments) and in vivo (anaesthetized animals with cannulation of the common bile duct) whereas the trophic effect was investigated after short-term (5 days) administration of the peptides and/or lorglumide. 3. Both caerulein and bombesin stimulated amylase release from in vitro pancreatic segments in a concentration-dependent manner. Although the efficacy of both peptides was virtually identical, the potency of caerulein was higher than that of bombesin. Lorglumide displaced the concentration-response curves to caerulein to the right without affecting the maximum response, suggesting a competitive antagonism. The Schild plot analysis of data gave a straight line with a slope not significantly different from unity. The calculated pA2 for lorglumide was 7.31 +/- 0.45. The antagonist, however, was completely ineffective when tested against bombesin-induced amylase release. 4. In vivo experiments confirmed results from in vitro studies since lorglumide (5 and 10 mg kg-1) significantly reduced pancreatic exocrine secretion induced by caerulein without affecting the response to bombesin. 5. Administration of either peptide increased the weight of the pancreas, the total pancreatic protein and DNA, trypsin and amylase content. Lorglumide (10 mg kg-1), administered together with caerulein, reduced the peptide-induced increase in pancreatic weight, protein and enzyme content. On the contrary, when lorglumide was given together with bombesin, all the parameters that were examined were not altered by concomitant administration of the antagonist. 6. These results have demonstrated the ability of lorglumide to antagonize the effects on the pancreas of a CCK-analogue, caerulein, and its inability to affect bombesin-induced

CHARACTERIZATION OF THE ALKANE-INDUCIBLE CYTOCHROME P450 (P450ALK) GENE FROM THE YEAST CANDIDA TROPICALIS: IDENTIFICATION OF A NEW P450 FAMILY

EPA Science Inventory

The P450alk gene, which is inducible by the assimilation of alkane in Candida tropicalis, was sequenced and characterized. Structural features described in promoter and terminator regions of Saccharomyces yeast genes are present in the P450alk gene and some particular structures ...

2',5'-Dihydroxychalcone as a potent chemical mediator and cyclooxygenase inhibitor.

PubMed

Lin, C N; Lee, T H; Hsu, M F; Wang, J P; Ko, F N; Teng, C M

1997-05-01

Eleven chalcone derivatives have been tested for their inhibitory effects on platelet aggregation in rabbit platelet suspension and the activation of mast cells and neutrophils. Arachidonic acid-induced platelet aggregation was potently inhibited by almost all the compounds and some also had a potent inhibitory effect on collagen-induced platelet aggregation and cyclooxygenase. Some hydroxychalcone derivatives showed strong inhibitory effects on the release of beta-glucuronidase and lysozyme, and on superoxide formation by rat neutrophils stimulated with the peptide fMet-Leu-Phe (fMLP). We found that the anti-inflammatory effect of 2',5'-dihydroxychalcone was greater than that of trifluoperazine. 2'5'-Dihydroxy and 2',3,4,5'-tetrahydroxyl chalcones, even at low concentration (50 microM), tested in platelet-rich plasma from man almost completely inhibited secondary aggregation induced by adrenaline. These results suggest that the anti-platelet effects of the chalcones are mainly a result of inhibition of thromboxane formation.

Inducible DNA-repair systems in yeast: competition for lesions.

PubMed

Mitchel, R E; Morrison, D P

1987-03-01

DNA lesions may be recognized and repaired by more than one DNA-repair process. If two repair systems with different error frequencies have overlapping lesion specificity and one or both is inducible, the resulting variable competition for the lesions can change the biological consequences of these lesions. This concept was demonstrated by observing mutation in yeast cells (Saccharomyces cerevisiae) exposed to combinations of mutagens under conditions which influenced the induction of error-free recombinational repair or error-prone repair. Total mutation frequency was reduced in a manner proportional to the dose of 60Co-gamma- or 254 nm UV radiation delivered prior to or subsequent to an MNNG exposure. Suppression was greater per unit radiation dose in cells gamma-irradiated in O2 as compared to N2. A rad3 (excision-repair) mutant gave results similar to wild-type but mutation in a rad52 (rec-) mutant exposed to MNNG was not suppressed by radiation. Protein-synthesis inhibition with heat shock or cycloheximide indicated that it was the mutation due to MNNG and not that due to radiation which had changed. These results indicate that MNNG lesions are recognized by both the recombinational repair system and the inducible error-prone system, but that gamma-radiation induction of error-free recombinational repair resulted in increased competition for the lesions, thereby reducing mutation. Similarly, gamma-radiation exposure resulted in a radiation dose-dependent reduction in mutation due to MNU, EMS, ENU and 8-MOP + UVA, but no reduction in mutation due to MMS. These results suggest that the number of mutational MMS lesions recognizable by the recombinational repair system must be very small relative to those produced by the other agents. MNNG induction of the inducible error-prone systems however, did not alter mutation frequencies due to ENU or MMS exposure but, in contrast to radiation, increased the mutagenic effectiveness of EMS. These experiments demonstrate

[Defects in TOR regulatory complexes retard aging and carbonyl/oxidative stress development in yeast Saccharomyces cerevisiae].

PubMed

Homza, B V; Vasyl'kovs'ka, R A; Semchyshyn, H M

2014-01-01

TOR signaling pathway first described in yeast S. cerevisiae is the highly conserved regulator of eukaryotic cell growth, aging and stress resistance. The effect of nitrogen sources, in particular amino acids, on the activity of TOR signaling pathway is well studied, however its relation to carbohydrates is poor understood. The aim of the present study is expanding of our understanding of potential role of TOR regulatory complexes in development of carbonyl/oxidative stress that can result from yeast cultivation on glucose and fructose. It has been shown that the level of alpha-dicarbonyl compounds and protein carbonyl groups increased with time of yeast cultivation and was higher in cells grown on fructose that demonstrated their accelerated aging and carbonyl/oxidative stress development as compared with cells grown on glucose. The strains defective in TOR proteins cultivated in the presence of glucose as well as fructose demonstrated lower markers of the stress and aging than parental strain. Thus these data confirmed the previous conclusion on fructose more potent ability to cause carbonyl/oxidative stress and accelerated aging in S. cerevisiae as compared with glucose. However, defects in TOR regulatory complexes retard aging and development of the stress in yeast independent on the type of carbohydrate in the cultivation medium.

Prevalence of HBsAg, knowledge, and vaccination practice against viral hepatitis B infection among doctors and nurses in a secondary health care facility in Lagos state, South-western Nigeria.

PubMed

Abiola, Abdul-Hakeem Olatunji; Agunbiade, Adebukola Bola; Badmos, Kabir Bolarinwa; Lesi, Adenike Olufunmilayo; Lawal, Abdulrazzaq Oluwagbemiga; Alli, Quadri Olatunji

2016-01-01

Hepatitis B Virus, a highly infectious blood-borne virus poses a major threat to public health globally due to its high prevalence rate and grave consequence in causing liver cirrhosis and hepatocelullar carcinoma, the third cause of cancer death worldwide. The aim is determine the prevalence of HBsAg, knowledge, and vaccination practices against viral hepatitis B infection among doctors and nurses in a health care facility. Study design was a descriptive cross-sectional study among all the doctors and nurses in the health care facility. Data was collected using pre-tested, structured, self-administered questionnaire and blood samples were taken from respondents and tested using commercial enzyme-linked immunosorbent assay (ELIZA) test kit to determine prevalence of hepatitis B surface antigen after informed consent. Ethical approval was obtained from Health Research and Ethics Committee of the Lagos University Teaching Hospital. Responses of the respondents to the knowledge and vaccination practices against viral hepatitis B infection were scored and graded as poor (<50%), fair (50-74%) and good (≥75%). The study was carried out in January, 2014. A total of 134 out of the 143 recruited respondents participated in the study. Prevalence of HBsAg was 1.5%. Among the respondents, 56.7% had good knowledge and 94.8% reported poor practice of vaccination against viral hepatitis B infection. Mean knowledge and vaccination practices scores (%) were 72.54+7.60 and 29.44+14.37 respectively. Only 29% of the respondents did post vaccination testing for anti HBsAg. Prevalence of HBsAg was low. Knowledge of viral hepatitis B was fair, and practice of post hepatitis B vaccination testing was poor. It is therefore recommended that the state ministry of health should organise further health education programme, institute compulsory occupational hepatitis B vaccination programme and post vaccination anti-HBS testing to ensure adequate antibody level in this adult population.

Glycine betaine improves oxidative stress tolerance and biocontrol efficacy of antagonistic yeast Cystofilobasidium infirmominiatum

USDA-ARS?s Scientific Manuscript database

The effect of H2O2-induced oxidative stress on the viability of the yeast antagonist, Cystofilobasidium infirmominiatum, as well as the effect of exogenous glycine betaine (GB) on yeast viability under oxidative stress, was determined. GB treatment improved the tolerance of C. infirmominiatum to ox...

Method for using a yeast alpha-amylase promoter

DOEpatents

Gao, Johnway; Skeen, Rodney S.; Hooker, Brian S.; Anderson, Daniel B.

2003-04-22

The present invention provides the promoter clone discovery of an alpha-amylase gene of a starch utilizing yeast strain Schwanniomyces castellii. The isolated alpha-amylase promoter is an inducible promoter, which can regulate strong gene expression in starch culture medium.

Dynamic of Immune Response induced in Hepatitis B Surface Antigen-transgenic Mice Immunized with a Novel Therapeutic Formulation

PubMed Central

Almeida, Freya M Freyre; Blanco, Aracelys; Trujillo, Heidy; Hernández, Dunia; García, Daymir; Alba, José S; Abad, Matilde López; Merino, Nelson; Lobaina, Yadira

2016-01-01

ABSTRACT The development of therapeutic vaccines against chronic hepatitis B requires the capacity of the formulation to subvert a tolerated immune response as well as the evaluation of histopathological damage resulting from the treatment. In the present study, the dynamicity of induced immune response to hepatitis B surface antigen (HBsAg) was evaluated in transgenic mice that constitutively express the HBsAg gene (HBsAg-tg mice). After immunization with a vaccine candidate containing both surface (HBsAg) and core (HBcAg) antigens of hepatitis B virus (HBV), the effect of vaccination on clearance of circulating HBsAg and the potential histological alterations were examined. Transgenic (tg) and non-transgenic (Ntg) mice were immunized by intranasal (IN) and subcutaneous (SC) routes simultaneously. A control group received phosphate-buffered saline (PBS) by IN route and aluminum by SC route. Positive responses, at both humoral and cellular levels, were obtained after five immunizations in HBsAg-tg mice. Such responses were delayed and of lower intensity in tg mice, compared to vaccinated Ntg mice. Serum IgG response was characterized by a similar IgG subclass pattern. Even when HBsAg-specific CD8+ T cell responses were clearly detectable by gamma-interferon ELISPOT assay, histopathological alterations were not detected in any organ, including the liver and kidneys. Our study demonstrated, that it is possible to subvert the immune tolerance against HBsAg in tg mice, opening a window for new studies to optimize the schedule, dose, and formulation to improve the immune response to the therapeutic vaccine candidate. These results can be considered a safety proof to support clinical developments for the formulation under study. How to cite this article Freyre FM, Blanco A, Trujillo H, Hernández D, García D, Alba JS, Lopez M, Merino N, Lobaina Y, Aguilar JC. Dynamic of Immune Response induced in Hepatitis B Surface Antigen-transgenic Mice Immunized with a Novel

Denatured states of yeast cytochrome c induced by heat and guanidinium chloride are structurally and thermodynamically different.

PubMed

Zaidi, Sobia; Haque, Md Anzarul; Ubaid-Ullah, Shah; Prakash, Amresh; Hassan, Md Imtaiyaz; Islam, Asimul; Batra, Janendra K; Ahmad, Faizan

2017-05-01

A sequence alignment of mammalian cytochromes c with yeast iso-1-cytochrome c (y-cyt-c) shows that the yeast protein contains five extra N-terminal residues. We have been interested in understanding the question: What is the role of these five extra N-terminal residues in folding and stability of the protein? To answer this question we have prepared five deletants of y-cyt-c by sequentially removing these extra residues. During our studies on the wild type (WT) protein and its deletants, we observed that the amount of secondary structure in the guanidinium chloride (GdmCl)-induced denatured (D) state of each protein is different from that of the heat-induced denatured (H) state. This finding is confirmed by the observation of an additional cooperative transition curve of optical properties between H and D states on the addition of different concentrations of GdmCl to the already heat denatured WT y-cyt-c and its deletants at pH 6.0 and 68°C. For each protein, analysis of transition curves representing processes, native (N) state ↔ D state, N state ↔ H state, and H state ↔ D state, was done to obtain Gibbs free energy changes associated with all the three processes. This analysis showed that, for each protein, thermodynamic cycle accommodates Gibbs free energies associated with transitions between N and D states, N and H states, and H and D states, the characteristics required for a thermodynamic function. All these experimental observations have been supported by our molecular dynamics simulation studies.

Structural differences between yeast and mammalian microtubules revealed by cryo-EM

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

Howes, Stuart C.; Geyer, Elisabeth A.; LaFrance, Benjamin

Microtubules are polymers of αβ-tubulin heterodimers essential for all eukaryotes. Despite sequence conservation, there are significant structural differences between microtubules assembled in vitro from mammalian or budding yeast tubulin. Yeast MTs were not observed to undergo compaction at the interdimer interface as seen for mammalian microtubules upon GTP hydrolysis. Lack of compaction might reflect slower GTP hydrolysis or a different degree of allosteric coupling in the lattice. The microtubule plus end–tracking protein Bim1 binds yeast microtubules both between αβ-tubulin heterodimers, as seen for other organisms, and within tubulin dimers, but binds mammalian tubulin only at interdimer contacts. At the concentrationsmore » used in cryo-electron microscopy, Bim1 causes the compaction of yeast microtubules and induces their rapid disassembly. In conclusion, our studies demonstrate structural differences between yeast and mammalian microtubules that likely underlie their differing polymerization dynamics. These differences may reflect adaptations to the demands of different cell size or range of physiological growth temperatures.« less

Forging a potent vaccine adjuvant: CpG ODN/cationic peptide nanorings.

PubMed

Gungor, Bilgi; Yagci, Fuat Cem; Gursel, Ihsan; Gursel, Mayda

Type I interferon inducers may potentially be engineered to function as antiviral and anticancer agents, or alternatively, vaccine adjuvants, all of which may have clinical applications. We recently described a simple strategy to convert a Toll-like receptor 9 (TLR9) agonist devoid of interferon α (IFNα) stimulating activity into a robust Type I interferon inducer with potent vaccine adjuvant activity.

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

Glycine betaine improves oxidative stress tolerance and biocontrol efficacy of the antagonistic yeast Cystofilobasidium infirmominiatum

USDA-ARS?s Scientific Manuscript database

The effect of hydrogen peroxide-induced oxidative stress on the viability of the yeast antagonist, Cystofilobasidium infirmominiatum, as well as the effect of exogenous glycine betaine (GB) on yeast viability under oxidative stress, was determined. GB treatment improved the tolerance of C. infirmom...

Drosophila Regulate Yeast Density and Increase Yeast Community Similarity in a Natural Substrate

PubMed Central

Stamps, Judy A.; Yang, Louie H.; Morales, Vanessa M.; Boundy-Mills, Kyria L.

2012-01-01

Drosophila melanogaster adults and larvae, but especially larvae, had profound effects on the densities and community structure of yeasts that developed in banana fruits. Pieces of fruit exposed to adult female flies previously fed fly-conditioned bananas developed higher yeast densities than pieces of the same fruits that were not exposed to flies, supporting previous suggestions that adult Drosophila vector yeasts to new substrates. However, larvae alone had dramatic effects on yeast density and species composition. When yeast densities were compared in pieces of the same fruits assigned to different treatments, fruits that developed low yeast densities in the absence of flies developed significantly higher yeast densities when exposed to larvae. Across all of the fruits, larvae regulated yeast densities within narrow limits, as compared to a much wider range of yeast densities that developed in pieces of the same fruits not exposed to flies. Larvae also affected yeast species composition, dramatically reducing species diversity across fruits, reducing variation in yeast communities from one fruit to the next (beta diversity), and encouraging the consistent development of a yeast community composed of three species of yeast (Candida californica, C. zemplinina, and Pichia kluvyeri), all of which were palatable to larvae. Larvae excreted viable cells of these three yeast species in their fecal pools, and discouraged the growth of filamentous fungi, processes which may have contributed to their effects on the yeast communities in banana fruits. These and other findings suggest that D. melanogaster adults and their larval offspring together engage in ‘niche construction’, facilitating a predictable microbial environment in the fruit substrates in which the larvae live and develop. PMID:22860093

A sulforaphane analogue that potently activates the Nrf2-dependent detoxification pathway.

PubMed

Morimitsu, Yasujiro; Nakagawa, Yoko; Hayashi, Kazuhiro; Fujii, Hiroyuki; Kumagai, Takeshi; Nakamura, Yoshimasa; Osawa, Toshihiko; Horio, Fumihiko; Itoh, Ken; Iida, Katsuyuki; Yamamoto, Masayuki; Uchida, Koji

2002-02-01

Exposure of cells to a wide variety of chemoprotective compounds confers resistance to a broad set of carcinogens. For a subset of the chemoprotective compounds, protection is generated by an increase in the abundance of the protective phase II detoxification enzymes, such as glutathione S-transferase (GST). We have recently developed a cell culture system, using rat liver epithelial RL 34 cells, that potently responds to the phenolic antioxidants resulting in the induction of GST activity (Kawamoto, Y., Nakamura, Y., Naito, Y., Torii, Y., Kumagai, T., Osawa, T., Ohigashi, H., Satoh, K., Imagawa, M., and Uchida, K. (2000) J. Biol. Chem. 275, 11291-11299.) In the present study, we investigated the phase II-inducing potency of an isothiocyanate compound in vitro and in vivo and examined a possible induction mechanism. Based on an extensive screening of vegetable extracts for GST inducer activity in RL34 cells, we found Japanese horseradish, wasabi (Wasabia japonica, syn. Eutrema wasabi), as the richest source and identified 6-methylsulfinylhexyl isothiocyanate (6-HITC), an analogue of sulforaphane (4-methylsulfinylbutyl isothiocyanate) isolated from broccoli, as the major GST inducer in wasabi. 6-HITC potently induced both class alpha GSTA1 and class pi GSTP1 isozymes in RL34 cells. In animal experiments, we found that 6-MSHI was rapidly absorbed into the body and induced hepatic phase II detoxification enzymes more potently than sulforaphane. The observations that (i) 6-HITC activated the antioxidant response element (ARE), (ii) 6-HITC induced nuclear localization of the transcription factor Nrf2 that binds to ARE, and (iii) the induction of phase II enzyme genes by 6-HITC was completely abrogated in the nrf2-deficient mice, suggest that 6-HITC is a potential activator of the Nrf2/ARE-dependent detoxification pathway.

Effect of a novel selective and potent phosphinic peptide inhibitor of endopeptidase 3.4.24.16 on neurotensin-induced analgesia and neuronal inactivation.

PubMed

Vincent, B; Jiracek, J; Noble, F; Loog, M; Roques, B; Dive, V; Vincent, J P; Checler, F

1997-06-01

1. We have examined a series of novel phosphinic peptides as putative potent and selective inhibitors of endopeptidase 3.4.24.16. 2. The most selective inhibitor, Pro-Phe-psi(PO2CH2)-Leu-Pro-NH2 displayed a Ki value of 12 nM towards endopeptidase 3.4.24.16 and was 5540 fold less potent on its related peptidase endopeptidase 3.4.24.15. Furthermore, this inhibitor was 12.5 less potent on angiotensin-converting enzyme and was unable to block endopeptidase 3.4.24.11, aminopeptidases B and M, dipeptidylaminopeptidase IV and proline endopeptidase. 3. The effect of Pro-Phe-psi(PO2CH2)-Leu-Pro-NH2, in vitro and in vivo, on neurotensin metabolism in the central nervous system was examined. 4. Pro-Phe-psi(PO2CHH2)-Leu-Pro-NH2 dose-dependently inhibited the formation of neurotensin 1-10 and concomittantly protected neurotensin from degradation by primary cultured neurones from mouse embryos. 5. Intracerebroventricular administration of Pro-Phe-psi(PO2CH2)-Leu-Pro-NH2 significantly potentiated the neurotensin-induced antinociception of mice in the hot plate test. 6. Altogether, our study has established Pro-Phe-psi(PO2CH2)-Leu-Pro-NH2 as a fully selective and highly potent inhibitor of endopeptidase 3.4.24.16 and demonstrates, for the first time, the contribution of this enzyme in the central metabolism of neurotensin.

Effect of a novel selective and potent phosphinic peptide inhibitor of endopeptidase 3.4.24.16 on neurotensin-induced analgesia and neuronal inactivation

PubMed Central

Vincent, Bruno; Jiracek, Jirì; Noble, Florence; Loog, Mart; Roques, Bernard; Dive, Vincent; Vincent, Jean-Pierre; Checler, Frédéric

1997-01-01

We have examined a series of novel phosphinic peptides as putative potent and selective inhibitors of endopeptidase 3.4.24.16. The most selective inhibitor, Pro-Phe-Ψ(PO2CH2)-Leu-Pro-NH2 displayed a Ki value of 12 nM towards endopeptidase 3.4.24.16 and was 5540 fold less potent on its related peptidase endopeptidase 3.4.24.15. Furthermore, this inhibitor was 12.5 less potent on angiotensin-converting enzyme and was unable to block endopeptidase 3.4.24.11, aminopeptidases B and M, dipeptidylaminopeptidase IV and proline endopeptidase. The effect of Pro-Phe-Ψ(PO2CH2)-Leu-Pro-NH2, in vitro and in vivo, on neurotensin metabolism in the central nervous system was examined. Pro-Phe-Ψ(PO2CHH2)-Leu-Pro-NH2 dose-dependently inhibited the formation of neurotensin 1-10 and concomittantly protected neurotensin from degradation by primary cultured neurones from mouse embryos. Intracerebroventricular administration of Pro-Phe-Ψ(PO2CH2)-Leu-Pro-NH2 significantly potentiated the neurotensin-induced antinociception of mice in the hot plate test. Altogether, our study has established Pro-Phe-Ψ(PO2CH2)-Leu-Pro-NH2 as a fully selective and highly potent inhibitor of endopeptidase 3.4.24.16 and demonstrates, for the first time, the contribution of this enzyme in the central metabolism of neurotensin. PMID:9208137

UV and arsenate toxicity: a specific and sensitive yeast bioluminescence assay.

PubMed

Bakhrat, Anya; Eltzov, Evgeni; Finkelstein, Yishay; Marks, Robert S; Raveh, Dina

2011-06-01

We describe a Saccharomyces cerevisiae bioluminescence assay for UV and arsenate in which bacterial luciferase genes are regulated by the promoter of the yeast gene, UFO1. UFO1 encodes the F-box subunit of the Skp1–Cdc53–F-box protein ubiquitin ligase complex and is induced by DNA damage and by arsenate. We engineered the UFO1 promoter into an existing yeast bioreporter that employs human genes for detection of steroid hormone-disrupting compounds in water bodies. Our analysis indicates that use of an endogenous yeast promoter in different mutant backgrounds allows discrimination between different environmental signals. The UFO1-engineered yeast give a robust bioluminescence response to UVB and can be used for evaluating UV protective sunscreens. They are also effective in detecting extremely low concentrations of arsenate, particularly in pdr5Δ mutants that lack a mechanism to extrude toxic chemicals; however, they do not respond to cadmium or mercury. Combined use of endogenous yeast promoter elements and mutants of stress response pathways may facilitate development of high-specificity yeast bioreporters able to discriminate between closely related chemicals present together in the environment.

In vivo direct patulin-induced fluidization of the plasma membrane of fission yeast Schizosaccharomyces pombe.

PubMed

Horváth, Eszter; Papp, Gábor; Belágyi, József; Gazdag, Zoltán; Vágvölgyi, Csaba; Pesti, Miklós

2010-07-01

Patulin is a toxic metabolite produced by various species of Penicillium, Aspergillus and Byssochlamys. In the present study, its effects on the plasma membrane of fission yeast Schizosaccharomyces pombe were investigated. The phase-transition temperature (G) of untreated cells, measured by electron paramagnetic resonance spectrometry proved to be 14.1 degrees C. Treatment of cells for 20 min with 50, 500, or 1000 microM patulin resulted in a decrease of the G value of the plasma membrane to 13.9, 10.1 or 8.7 degrees C, respectively. This change in the transition temperature was accompanied by the loss of compounds absorbing light at 260 nm. Treatment of cells with 50, 500 or 1000 microM patulin for 20 min induced the efflux of 25%, 30.5% or 34%, respectively, of these compounds. Besides its cytotoxic effects an adaptation process was observed. This is the first study to describe the direct interaction of patulin with the plasma membrane, a process which could definitely contribute to the adverse toxic effects induced by patulin. 2010 Elsevier Ltd. All rights reserved.

Prion-based memory of heat stress in yeast

PubMed Central

Chernova, Tatiana A.; Wilkinson, Keith D.

2017-01-01

ABSTRACT Amyloids and amyloid-based prions are self-perpetuating protein aggregates which can spread by converting a normal protein of the same sequence into a prion form. They are associated with diseases in humans and mammals, and control heritable traits in yeast and other fungi. Some amyloids are implicated in biologically beneficial processes. As prion formation generates reproducible memory of a conformational change, prions can be considered as molecular memory devices.  We have demonstrated that in yeast, stress-inducible cytoskeleton-associated protein Lsb2 forms a metastable prion in response to high temperature. This prion promotes conversion of other proteins into prions and can persist in a fraction of cells for a significant number of cell generations after stress, thus maintaining the memory of stress in a population of surviving cells. Acquisition of an amino acid substitution required for Lsb2 to form a prion coincides with acquisition of increased thermotolerance in the evolution of Saccharomyces yeast. Thus the ability to form an Lsb2 prion in response to stress coincides with yeast adaptation to growth at higher temperatures. These findings intimately connect prion formation to the cellular response to environmental stresses. PMID:28521568

Prion-based memory of heat stress in yeast.

PubMed

Chernova, Tatiana A; Chernoff, Yury O; Wilkinson, Keith D

2017-05-04

Amyloids and amyloid-based prions are self-perpetuating protein aggregates which can spread by converting a normal protein of the same sequence into a prion form. They are associated with diseases in humans and mammals, and control heritable traits in yeast and other fungi. Some amyloids are implicated in biologically beneficial processes. As prion formation generates reproducible memory of a conformational change, prions can be considered as molecular memory devices.  We have demonstrated that in yeast, stress-inducible cytoskeleton-associated protein Lsb2 forms a metastable prion in response to high temperature. This prion promotes conversion of other proteins into prions and can persist in a fraction of cells for a significant number of cell generations after stress, thus maintaining the memory of stress in a population of surviving cells. Acquisition of an amino acid substitution required for Lsb2 to form a prion coincides with acquisition of increased thermotolerance in the evolution of Saccharomyces yeast. Thus the ability to form an Lsb2 prion in response to stress coincides with yeast adaptation to growth at higher temperatures. These findings intimately connect prion formation to the cellular response to environmental stresses.

Crossflow microfiltration of yeast suspensions in tubular filters.

PubMed

Redkar, S G; Davis, R H

1993-01-01

Crossflow microfiltration experiments were performed on yeast suspensions through 0.2-microns pore size ceramic and polypropylene tubes at various operating conditions. The initial transient flux decline follows dead-end filtration theory, with the membrane resistance determined from the initial flux and the specific cake resistance determined from the rate of flux decline due to cake buildup. For long times, the observed fluxes reach steady or nearly steady values, presumably as a result of the cake growth being arrested by the shear exerted at its surface. The steady-state fluxes increase with increasing shear rate and decreasing feed concentration, and they are nearly independent of transmembrane pressure. The steady-state fluxes for unwashed yeast in deionized water or fermentation media are typically 2-4 times lower than those predicted by a model based on the properties of nonadhesive, rigid spheres undergoing shear-induced back-diffusion. In contrast, the steady-state fluxes observed for washed yeast cells in deionized water are only 10-30% below the predicted values. The washed yeast cells also exhibited specific cake resistances that are an order of magnitude lower than those for the unwashed yeast. The differences are due to the presence of extracellular proteins and other macromolecules in the unwashed yeast suspensions. These biopolymers cause higher cell adhesion and resistance in the cake layer, so that the cells at the top edge are not free to diffuse away. This is manifested as a concentration jump from the edge of the cake layer to the sheared suspension adjacent to it.(ABSTRACT TRUNCATED AT 250 WORDS)

Therapeutic activity of a Saccharomyces cerevisiae-based probiotic and inactivated whole yeast on vaginal candidiasis

PubMed Central

Pericolini, Eva; Gabrielli, Elena; Ballet, Nathalie; Sabbatini, Samuele; Roselletti, Elena; Cayzeele Decherf, Amélie; Pélerin, Fanny; Luciano, Eugenio; Perito, Stefano; Jüsten, Peter; Vecchiarelli, Anna

2017-01-01

ABSTRACT Vulvovaginal candidiasis is the most prevalent vaginal infection worldwide and Candida albicans is its major agent. Vulvovaginal candidiasis is characterized by disruption of the vaginal microbiota composition, as happens following large spectrum antibiotic usage. Recent studies support the effectiveness of oral and local probiotic treatment for prevention of recurrent vulvovaginal candidiasis. Saccharomyces cerevisiae is a safe yeast used as, or for, the production of ingredients for human nutrition and health. Here, we demonstrate that vaginal administration of probiotic Saccharomyces cerevisiae live yeast (GI) and, in part, inactivated whole yeast Saccharomyces cerevisiae (IY), used as post-challenge therapeutics, was able to positively influence the course of vaginal candidiasis by accelerating the clearance of the fungus. This effect was likely due to multiple interactions of Saccharomyces cerevisiae with Candida albicans. Both live and inactivated yeasts induced coaggregation of Candida and consequently inhibited its adherence to epithelial cells. However, only the probiotic yeast was able to suppress some major virulence factors of Candida albicans such as the ability to switch from yeast to mycelial form and the capacity to express several aspartyl proteases. The effectiveness of live yeast was higher than that of inactivated whole yeast suggesting that the synergy between mechanical effects and biological effects were dominant over purely mechanical effects. The protection of epithelial cells to Candida-induced damage was also observed. Overall, our data show for the first time that Saccharomyces cerevisiae-based ingredients, particularly the living cells, can exert beneficial therapeutic effects on a widespread vaginal mucosal infection. PMID:27435998

Therapeutic activity of a Saccharomyces cerevisiae-based probiotic and inactivated whole yeast on vaginal candidiasis.

PubMed

Pericolini, Eva; Gabrielli, Elena; Ballet, Nathalie; Sabbatini, Samuele; Roselletti, Elena; Cayzeele Decherf, Amélie; Pélerin, Fanny; Luciano, Eugenio; Perito, Stefano; Jüsten, Peter; Vecchiarelli, Anna

2017-01-02

Vulvovaginal candidiasis is the most prevalent vaginal infection worldwide and Candida albicans is its major agent. Vulvovaginal candidiasis is characterized by disruption of the vaginal microbiota composition, as happens following large spectrum antibiotic usage. Recent studies support the effectiveness of oral and local probiotic treatment for prevention of recurrent vulvovaginal candidiasis. Saccharomyces cerevisiae is a safe yeast used as, or for, the production of ingredients for human nutrition and health. Here, we demonstrate that vaginal administration of probiotic Saccharomyces cerevisiae live yeast (GI) and, in part, inactivated whole yeast Saccharomyces cerevisiae (IY), used as post-challenge therapeutics, was able to positively influence the course of vaginal candidiasis by accelerating the clearance of the fungus. This effect was likely due to multiple interactions of Saccharomyces cerevisiae with Candida albicans. Both live and inactivated yeasts induced coaggregation of Candida and consequently inhibited its adherence to epithelial cells. However, only the probiotic yeast was able to suppress some major virulence factors of Candida albicans such as the ability to switch from yeast to mycelial form and the capacity to express several aspartyl proteases. The effectiveness of live yeast was higher than that of inactivated whole yeast suggesting that the synergy between mechanical effects and biological effects were dominant over purely mechanical effects. The protection of epithelial cells to Candida-induced damage was also observed. Overall, our data show for the first time that Saccharomyces cerevisiae-based ingredients, particularly the living cells, can exert beneficial therapeutic effects on a widespread vaginal mucosal infection.

Prions in Yeast

PubMed Central

Liebman, Susan W.; Chernoff, Yury O.

2012-01-01

The concept of a prion as an infectious self-propagating protein isoform was initially proposed to explain certain mammalian diseases. It is now clear that yeast also has heritable elements transmitted via protein. Indeed, the “protein only” model of prion transmission was first proven using a yeast prion. Typically, known prions are ordered cross-β aggregates (amyloids). Recently, there has been an explosion in the number of recognized prions in yeast. Yeast continues to lead the way in understanding cellular control of prion propagation, prion structure, mechanisms of de novo prion formation, specificity of prion transmission, and the biological roles of prions. This review summarizes what has been learned from yeast prions. PMID:22879407

Potent anti-seizure effects of D-leucine

PubMed Central

Hartman, Adam L.; Santos, Polan; O’Riordan, Kenneth J.; Stafstrom, Carl E.; Hardwick, J. Marie

2015-01-01

There are no effective treatments for millions of patients with intractable epilepsy. High-fat ketogenic diets may provide significant clinical benefit but are challenging to implement. Low carbohydrate levels appear to be essential for the ketogenic diet to work, but the active ingredients in dietary interventions remain elusive, and a role for ketogenesis has been challenged. A potential antiseizure role of dietary protein or of individual amino acids in the ketogenic diet is understudied. We investigated the two exclusively ketogenic amino acids, L-leucine and L-lysine, and found that only L-leucine potently protects mice when administered prior to the onset of seizures induced by kainic acid injection, but not by inducing ketosis. Unexpectedly, the D-enantiomer of leucine, which is found in trace amounts in the brain, worked as well or better than L-leucine against both kainic acid and 6 Hz electroshock-induced seizures. However, unlike L-leucine, D-leucine potently terminated seizures even after the onset of seizure activity. Furthermore, D-leucine, but not L-leucine, reduced long-term potentiation but had no effect on basal synaptic transmission in vitro. In a screen of candidate neuronal receptors, D-leucine failed to compete for binding by cognate ligands, potentially suggesting a novel target. Even at low doses, D-leucine suppressed ongoing seizures at least as effectively as diazepam but without sedative effects. These studies raise the possibility that D-leucine may represent a new class of anti-seizure agents, and that D-leucine may have a previously unknown function in eukaryotes. PMID:26054437

NAD(P)H-dependent aldose reductase from the xylose-assimilating yeast Candida tenuis. Isolation, characterization and biochemical properties of the enzyme.

PubMed Central

Neuhauser, W; Haltrich, D; Kulbe, K D; Nidetzky, B

1997-01-01

During growth on d-xylose the yeast Candida tenuis produces one aldose reductase that is active with both NADPH and NADH as coenzyme. This enzyme has been isolated by dye ligand and anion-exchange chromatography in yields of 76%. Aldose reductase consists ofa single 43 kDa polypeptide with an isoelectric point of 4.70. Initial velocity, product inhibition and binding studies are consistent with a compulsory-ordered, ternary-complex mechanism with coenzyme binding first and leaving last. The catalytic efficiency (kcat/Km) in d-xylose reduction at pH 7 is more than 60-fold higher than that in xylitol oxidation and reflects significant differences in the corresponding catalytic centre activities as well as apparent substrate-binding constants. The enzyme prefers NADP(H) approx. 2-fold to NAD(H), which is largely due to better apparent binding of the phosphorylated form of the coenzyme. NADP+ is a potent competitive inhibitor of the NADH-linked aldehyde reduction (Ki 1.5 microM), whereas NAD+ is not. Unlike mammalian aldose reductase, the enzyme from C. tenuis is not subject to oxidation-induced activation. Evidence of an essential lysine residue located in or near the coenzyme binding site has been obtained from chemical modification of aldose reductase with pyridoxal 5'-phosphate. The results are discussed in the context of a comparison of the enzymic properties of yeast and mammalian aldose reductase. PMID:9307017

Mitochondrial metabolism and stress response of yeast: Applications in fermentation technologies.

PubMed

Kitagaki, Hiroshi; Takagi, Hiroshi

2014-04-01

Mitochondria are sites of oxidative respiration. During sake brewing, sake yeasts are exposed to long periods of hypoxia; the structure, role, and metabolism of mitochondria of sake yeasts have not been studied in detail. It was first elucidated that the mitochondrial structure of sake yeast transforms from filamentous to dotted structure during sake brewing, which affects malate metabolism. Based on the information of yeast mitochondria during sake brewing, practical technologies have been developed; (i) breeding pyruvate-underproducing sake yeast by the isolation of a mutant resistant to an inhibitor of mitochondrial pyruvate transport; and (ii) modifying malate and succinate production by manipulating mitochondrial activity. During the bread-making process, baker's yeast cells are exposed to a variety of baking-associated stresses, such as freeze-thaw, air-drying, and high sucrose concentrations. These treatments induce oxidative stress generating reactive oxygen species due to mitochondrial damage. A novel metabolism of proline and arginine catalyzed by N-acetyltransferase Mpr1 in the mitochondria eventually leads to synthesis of nitric oxide, which confers oxidative stress tolerance on yeast cells. The enhancement of proline and arginine metabolism could be promising for breeding novel baker's yeast strains that are tolerant to multiple baking-associated stresses. These new and practical methods provide approaches to improve the processes in the field of industrial fermentation technologies. Copyright © 2013 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Characterization of the interaction of yeast enolase with polynucleotides.

PubMed

al-Giery, A G; Brewer, J M

1992-09-23

Yeast enolase is inhibited under certain conditions by DNA. The enzyme binds to single-stranded DNA-cellulose. Inhibition was used for routine characterization of the interaction. The presence of the substrate 2-phospho-D-glycerate reduces inhibition and binding. Both yeast enolase isozymes behave similarly. Impure yeast enolase was purified by adsorption onto a single-stranded DNA-cellulose column followed by elution with substrate. Interaction with RNA, double-stranded DNA, or degraded DNA results in less inhibition, suggesting that yeast enolase preferentially binds single-stranded DNA. However, yeast enolase is not a DNA-unwinding protein. The enzyme is inhibited by the short synthetic oligodeoxynucleotides G6, G8 and G10 but not T8 or T6, suggesting some base specificity in the interaction. The interaction is stronger at more acid pH values, with an apparent pK of 5.6. The interaction is prevented by 0.3 M KCl, suggesting that electrostatic factors are important. Histidine or lysine reverse the inhibition at lower concentrations, while phosphate is still more effective. Binding of single-stranded DNA to enolase reduces the reaction of protein histidyl residues with diethylpyrocarbonate. The inhibition of yeast enolase by single-stranded DNA is not total, and suggests the active site is not directly involved in the interaction. Binding of substrate may induce a conformational change in the enzyme that interferes with DNA binding and vice versa.

DNA Polymerase ζ is essential for hexavalent chromium-induced mutagenesis

PubMed Central

O'Brien, Travis J.; Witcher, Preston; Brooks, Bradford; Patierno, Steven R.

2009-01-01

Translesion synthesis (TLS) is a unique DNA damage tolerance mechanism involved in the replicative bypass of genetic lesions in favor of uninterrupted DNA replication. TLS is critical for the generation of mutations by many different chemical and physical agents, however, there is no information available regarding the role of TLS in carcinogenic metal-induced mutagenesis. Hexavalent chromium (Cr(VI))-containing compounds are highly complex genotoxins possessing both mutagenic and clastogenic activities. The focus of this work was to determine the impact that TLS has on Cr(VI)-induced mutagenesis in S. cerevisiae. Wild-type yeast and strains deficient in TLS polymerases (i.e. Polζ (rev3), Polη (rad30)) were exposed to Cr(VI) and monitored for cell survival and forward mutagenesis at the CAN1 locus. In general, TLS deficiency had little impact on Cr(VI)-induced clonogenic lethality or cell growth. rad30 yeast exhibited higher levels of basal and induced mutagenesis compared to Wt and rev3 yeast. In contrast, rev3 yeast displayed attenuated Cr(VI)-induced mutagenesis. Moreover, deletion of REV3 in rad30 yeast (rad30 rev3) resulted in a significant decrease in basal and Cr(VI) mutagenesis relative to Wt and rad30 single mutants indicating that mutagenesis primarily depended upon Polζ. Interestingly, rev3 yeast were similar to Wt yeast in susceptibility to Cr(VI)-induced frameshift mutations. Mutational analysis of the CAN1 gene revealed that Cr(VI)-induced base substitution mutations accounted for 83.9% and 100.0% of the total mutations in Wt and rev3 yeast, respectively. Insertions and deletions comprised 16.1% of the total mutations in Cr(VI) treated Wt yeast but were not observed rev3 yeast. This work provides novel information regarding the molecular mechanisms of Cr(VI)-induced mutagenesis and is the first report demonstrating a role for TLS in the fixation of mutations induced by a carcinogenic metal. PMID:19428373

Visible light alters yeast metabolic rhythms by inhibiting respiration.

PubMed

Robertson, James Brian; Davis, Chris R; Johnson, Carl Hirschie

2013-12-24

Exposure of cells to visible light in nature or in fluorescence microscopy often is considered to be relatively innocuous. However, using the yeast respiratory oscillation (YRO) as a sensitive measurement of metabolism, we find that non-UV visible light has a significant impact on yeast metabolism. Blue/green wavelengths of visible light shorten the period and dampen the amplitude of the YRO, which is an ultradian rhythm of cell metabolism and transcription. The wavelengths of light that have the greatest effect coincide with the peak absorption regions of cytochromes. Moreover, treating yeast with the electron transport inhibitor sodium azide has similar effects on the YRO as visible light. Because impairment of respiration by light would change several state variables believed to play vital roles in the YRO (e.g., oxygen tension and ATP levels), we tested oxygen's role in YRO stability and found that externally induced oxygen depletion can reset the phase of the oscillation, demonstrating that respiratory capacity plays a role in the oscillation's period and phase. Light-induced damage to the cytochromes also produces reactive oxygen species that up-regulate the oxidative stress response gene TRX2 that is involved in pathways that enable sustained growth in bright visible light. Therefore, visible light can modulate cellular rhythmicity and metabolism through unexpectedly photosensitive pathways.

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

Miltefosine is fungicidal to Paracoccidioides spp. yeast cells but subinhibitory concentrations induce melanisation.

PubMed

Rossi, Diego Conrado Pereira; Spadari, Cristina de Castro; Nosanchuk, Joshua Daniel; Taborda, Carlos Pelleschi; Ishida, Kelly

2017-04-01

Paracoccidioidomycosis (PCM) is a systemic mycosis caused by the dimorphic fungi Paracoccidioides spp. The duration of antifungal treatment ranges from months to years and relapses may nevertheless occur despite protracted therapy. Thus, there remains an urgent need for new therapeutic options. Miltefosine (MLT), an analogue of alkylphospholipids, has antifungal activity against species of yeast and filamentous fungi. The aim of this study was to evaluate the antifungal effects of MLT on the yeast forms of Paracoccidioides brasiliensis and Paracoccidioides lutzii. MLT demonstrated inhibitory activity from 0.12 to 1 µg/mL, which was similar to amphotericin B or the combination trimethoprim/sulfamethoxazole but was not more effective than itraconazole. The fungicidal activity of MLT occurred at concentrations ≥1 µg/mL. Ultrastructural alterations were observed following exposure of the fungus to a subinhibitory concentration of MLT, such as cytoplasmic membrane alteration, mitochondrial swelling, electron-lucent vacuole accumulation and increasing melanosome-like structures. Melanin production by yeasts following MLT exposure was confirmed by labelling with antibodies to melanin. In addition, the combination of a subinhibitory concentration of MLT and tricyclazole, an inhibitor of DHN-melanin biosynthesis, drastically reduced yeast viability. In conclusion, MLT had a fungicidal effect against both Paracoccidioides spp., and a subinhibitory concentration impacted melanogenesis. These findings suggest that additional investigations should be pursued to establish a role for MLT in the treatment of PCM. Copyright © 2017 Elsevier B.V. and International Society of Chemotherapy. All rights reserved.

How does yeast respond to pressure?

PubMed

Fernandes, P M B

2005-08-01

The brewing and baking yeast Saccharomyces cerevisiae has been used as a model for stress response studies of eukaryotic cells. In this review we focus on the effect of high hydrostatic pressure (HHP) on S. cerevisiae. HHP exerts a broad effect on yeast cells characteristic of common stresses, mainly associated with protein alteration and lipid bilayer phase transition. Like most stresses, pressure induces cell cycle arrest. Below 50 MPa (500 atm) yeast cell morphology is unaffected whereas above 220 MPa wild-type cells are killed. S. cerevisiae cells can acquire barotolerance if they are pretreated with a sublethal stress due to temperature, ethanol, hydrogen peroxide, or pressure. Nevertheless, pressure only leads to protection against severe stress if, after pressure pretreatment, the cells are also re-incubated at room pressure. We attribute this effect to the inhibition of the protein synthesis apparatus under HHP. The global genome expression analysis of S. cerevisiae cells submitted to HHP revealed a stress response profile. The majority of the up-regulated genes are involved in stress defense and carbohydrate metabolism while most repressed genes belong to the cell cycle progression and protein synthesis categories. However, the signaling pathway involved in the pressure response is still to be elucidated. Nitric oxide, a signaling molecule involved in the regulation of a large number of cellular functions, confers baroprotection. Furthermore, S. cerevisiae cells in the early exponential phase submitted to 50-MPa pressure show induction of the expression level of the nitric oxide synthase inducible isoform. As pressure becomes an important biotechnological tool, studies concerning this kind of stress in microorganisms are imperative.

Yeast nitrogen utilization in the phyllosphere during plant lifespan under regulation of autophagy

PubMed Central

Shiraishi, Kosuke; Oku, Masahide; Kawaguchi, Kosuke; Uchida, Daichi; Yurimoto, Hiroya; Sakai, Yasuyoshi

2015-01-01

Recently, microbe-plant interactions at the above-ground parts have attracted great attention. Here we describe nitrogen metabolism and regulation of autophagy in the methylotrophic yeast Candida boidinii, proliferating and surviving on the leaves of Arabidopsis thaliana. After quantitative analyses of yeast growth on the leaves of A. thaliana with the wild-type and several mutant yeast strains, we showed that on young leaves, nitrate reductase (Ynr1) was necessary for yeast proliferation, and the yeast utilized nitrate as nitrogen source. On the other hand, a newly developed methylamine sensor revealed appearance of methylamine on older leaves, and methylamine metabolism was induced in C. boidinii, and Ynr1 was subjected to degradation. Biochemical and microscopic analysis of Ynr1 in vitro during a shift of nitrogen source from nitrate to methylamine revealed that Ynr1 was transported to the vacuole being the cargo for biosynthetic cytoplasm-to-vacuole targeting (Cvt) pathway, and degraded. Our results reveal changes in the nitrogen source composition for phyllospheric yeasts during plant aging, and subsequent adaptation of the yeasts to this environmental change mediated by regulation of autophagy. PMID:25900611

L-arabinose fermenting yeast

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

Zhang, Min; Singh, Arjun; Suominen, Pirkko

An L-arabinose utilizing yeast strain is provided for the production of ethanol by introducing and expressing bacterial araA, araB and araD genes. L-arabinose transporters are also introduced into the yeast to enhance the uptake of arabinose. The yeast carries additional genomic mutations enabling it to consume L-arabinose, even as the only carbon source, and to produce ethanol. A yeast strain engineered to metabolize arabinose through a novel pathway is also disclosed. Methods of producing ethanol include utilizing these modified yeast strains.

L-arabinose fermenting yeast

DOEpatents

Zhang, Min; Singh, Arjun; Suominen, Pirkko; Knoshaug, Eric; Franden, Mary Ann; Jarvis, Eric

2014-09-23

An L-arabinose utilizing yeast strain is provided for the production of ethanol by introducing and expressing bacterial araA, araB and araD genes. L-arabinose transporters are also introduced into the yeast to enhance the uptake of arabinose. The yeast carries additional genomic mutations enabling it to consume L-arabinose, even as the only carbon source, and to produce ethanol. A yeast strain engineered to metabolize arabinose through a novel pathway is also disclosed. Methods of producing ethanol include utilizing these modified yeast strains.

Resveratrol suppresses ethanol stress in winery and bottom brewery yeast by affecting superoxide dismutase, lipid peroxidation and fatty acid profile.

PubMed

Gharwalova, Lucia; Sigler, Karel; Dolezalova, Jana; Masak, Jan; Rezanka, Tomas; Kolouchova, Irena

2017-11-03

Mid-exponential cultures of two traditional biotechnological yeast species, winery Saccharomyces cerevisiae and the less ethanol tolerant bottom-fermenting brewery Saccharomyces pastorianus, were exposed to different concentrations of added ethanol (3, 5 and 8%) The degree of ethanol-induced cell stress was assessed by measuring the cellular activity of superoxide dismutase (SOD), level of lipid peroxidation products, changes in cell lipid content and fatty acid profile. The resveratrol as an antioxidant was found to decrease the ethanol-induced rise of SOD activity and suppress the ethanol-induced decrease in cell lipids. A lower resveratrol concentration (0.5 mg/l) even reduced the extent of lipid peroxidation in cells. Resveratrol also alleviated ethanol-induced changes in cell lipid composition in both species by strongly enhancing the proportion of saturated fatty acids and contributing thereby to membrane stabilization. Lower resveratrol concentrations could thus diminish the negative effects of ethanol stress on yeast cells and improve their physiological state. These effects may be utilized to enhance yeast vitality in high-ethanol-producing fermentations or to increase the number of yeast generations in brewery.

Prevention of Yeast Spoilage in Feed and Food by the Yeast Mycocin HMK

PubMed Central

Lowes, K. F.; Shearman, C. A.; Payne, J.; MacKenzie, D.; Archer, D. B.; Merry, R. J.; Gasson, M. J.

2000-01-01

The yeast Williopsis mrakii produces a mycocin or yeast killer toxin designated HMK; this toxin exhibits high thermal stability, high pH stability, and a broad spectrum of activity against other yeasts. We describe construction of a synthetic gene for mycocin HMK and heterologous expression of this toxin in Aspergillus niger. Mycocin HMK was fused to a glucoamylase protein carrier, which resulted in secretion of biologically active mycocin into the culture media. A partial purification protocol was developed, and a comparison with native W. mrakii mycocin showed that the heterologously expressed mycocin had similar physiological properties and an almost identical spectrum of biological activity against a number of yeasts isolated from silage and yoghurt. Two food and feed production systems prone to yeast spoilage were used as models to assess the ability of mycocin HMK to act as a biocontrol agent. The onset of aerobic spoilage in mature maize silage was delayed by application of A. niger mycocin HMK on opening because the toxin inhibited growth of the indigenous spoilage yeasts. This helped maintain both higher lactic acid levels and a lower pH. In yoghurt spiked with dairy spoilage yeasts, A. niger mycocin HMK was active at all of the storage temperatures tested at which yeast growth occurred, and there was no resurgence of resistant yeasts. The higher the yeast growth rate, the more effective the killing action of the mycocin. Thus, mycocin HMK has potential applications in controlling both silage spoilage and yoghurt spoilage caused by yeasts. PMID:10698773

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

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

Identification of a Potent Tryptophan-based TRPM8 Antagonist With in vivo Analgesic Activity.

PubMed

Bertamino, Alessia; Iraci, Nunzio; Ostacolo, Carmine; Ambrosino, Paolo; Musella, Simona; Di Sarno, Veronica; Ciaglia, Tania; Pepe, Giacomo; Sala, Marina; Soldovieri, Maria Virginia; Mosca, Ilaria; Gonzalez-Rodriguez, Sara; Fernández-Carvajal, Asia; Ferrer-Montiel, Antonio; Novellino, Ettore; Taglialatela, Maurizio; Campiglia, Pietro; Gomez-Monterrey, Isabel M

2018-06-25

TRPM8 has been implicated in nociception and pain and is currently regarded as an attractive target for the pharmacological treatment of neuropathic pain syndromes. A series of analogues of N,N'-dibenzyl tryptamine 1, a potent TRPM8 antagonist, were prepared and screened using a fluorescence-based in vitro assay based on menthol-evoked calcium influx in TRPM8 stably-transfected HEK293 cells. The tryptophan derivative 14 was identified as a potent (IC 50 0.2±0.2 nM) and selective TRPM8 antagonist. In vivo, 14 showed significant target coverage in both an icilin-induced WDS (at 1-30 mg/kg s.c.) and oxaliplatin-induced cold allodynia (at 0.1-1 μg s.c.) mice models. Molecular modeling studies identified the putative binding mode of these antagonists, suggesting that they could influence an interaction network between the S1-4 transmembrane segments and the TRP domains of the channel subunits. The tryptophan moiety provides a new pharmacophoric scaffold for the design of highly potent modulators of TRPM8-mediated pain.

Mechanism of action of coumarin and silver(I)-coumarin complexes against the pathogenic yeast Candida albicans.

PubMed

Thati, Bhumika; Noble, Andy; Rowan, Raymond; Creaven, Bernadette S; Walsh, Maureen; McCann, Malachy; Egan, Denise; Kavanagh, Kevin

2007-08-01

The anti-fungal activity and mode of action of a range of silver(I)-coumarin complexes was examined. The most potent silver(I)-coumarin complexes, namely 7-hydroxycoumarin-3-carboxylatosilver(I), 6-hydroxycoumarin-3-carboxylatosilver(I) and 4-oxy-3-nitrocoumarinbis(1,10-phenanthroline)silver(I), had MIC80 values of between 69.1 and 4.6 microM against the pathogenic yeast Candida albicans. These compounds also reduced respiration, lowered the ergosterol content of cells and increased the trans-membrane leakage of amino acids. A number of the complexes disrupted cytochrome synthesis in the cell and induced the appearance of morphological features consistent with cell death by apoptosis. These compounds appear to act by disrupting the synthesis of cytochromes which directly affects the cell's ability to respire. A reduction in respiration leads to a depletion in ergosterol biosynthesis and a consequent disruption of the integrity of the cell membrane. Disruption of cytochrome biosynthesis may induce the onset of apoptosis which has been shown previously to be triggered by alteration in the location of cytochrome c. Silver(I)-coumarin complexes demonstrate good anti-fungal activity and manifest a mode of action distinct to that of the conventional azole and polyene drugs thus raising the possibility of their use when resistance to conventional drug has emerged or in combination with such drugs.

Yeast caspase-dependent apoptosis in Saccharomyces cerevisiae BY4742 induced by antifungal and potential antitumor agent clotrimazole.

PubMed

Kavakçıoğlu, Berna; Tarhan, Leman

2018-01-01

Clotrimazole is an antifungal medication commonly used in the treatment of fungal infections. There is also promising research on using clotrimazole against other diseases such as malaria, beriberi, tineapedis and cancer. It was aimed to investigate the apoptotic phenotype in Saccharomyces cerevisiae induced by clotrimazole. The exposure of S. cerevisiae to 10 µM clotrimazole for 3, 6 and 9 h caused to decrease in cell viability by 24.82 ± 0.81, 56.00 ± 1.54 and 77.59 ± 0.53%, respectively. It was shown by Annexin V-PI assay that 110 µM clotrimazole treatment caused to death by 35.5 ± 2.48% apoptotic and only 13.1 ± 0.08% necrotic pathway within 30 min. The occurrence of DNA strand breaks and condensation could be visualised by the TUNEL and DAPI stainings, respectively. Yeast caspase activity was induced 12.34 ± 0.71-fold after 110 µM clotrimazole treatment for 30 min compared to the control. The dependency of clotrimazole-induced apoptosis to caspase was also shown using Δyca1 mutant.

[Mutants of the yeast Saccharomyces cerevisiae characterized by enhanced induced mutagenesis. III. Effect of the him mutation on the effectiveness and specificity of UF-induced mutagenesis].

PubMed

Ivanov, E L; Koval'tsova, S V; Korolev, V G

1987-09-01

We have studied the influence of him1-1, him2-1, him3-1 and himX mutations on induction frequency and specificity of UV-induced adenine-dependent mutations in the yeast Saccharomyces cerevisiae. Him mutations do not render haploid cells more sensitive to the lethal action of UV-light; however, in him strains adenine-dependent mutations (ade1, ade2) were induced more frequently (1.5--2-fold), as compared to the HIM strain. An analysis of the molecular nature of ade2 mutants revealed that him1-1, him2-1 and himX mutations increase specifically the yield of transitions (AT----GC and GC----AT), whereas in the him3-1 strain the yield of transversions was enhanced as well. We suggest him mutations analysed to affect specific repair pathway for mismatch correction.

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.

Novel depsides as potential anti-inflammatory agents with potent inhibitory activity against Escherichia coli-induced interleukin-8 production.

PubMed

Lv, Peng-Cheng; Xiong, Jing; Chen, Jin; Wang, Kai-Rui; Mao, Wen-Jun; Zhu, Hai-Liang

2010-08-01

Sixteen novel depsides were synthesized for the first time. Their chemical structures were clearly determined by (1)H NMR, ESI mass spectra, and elemental analyses. All the compounds were assayed for antibacterial activities against three Gram-positive bacterial strains (Bacillus subtilis ATCC 6633, Staphylococcus aureus ATCC 6538, and Streptococcus faecalis ATCC 9790) and three Gram-negative bacterial strains (Escherichia coli ATCC 35218, Pseudomonas aeruginosa ATCC 13525, and Enterobacter cloacae ATCC 13047) by the MTT method. Compound 2-(2-methoxy-2-oxoethyl)phenyl 5-bromonicotinate (5) exhibited significant antibacterial activities against E. coli ATCC 35218 with an MIC of 0.78 microg/mL, which was superior to the positive control kanamycin B. In addition, compound 5 showed potent inhibitory activity against E. coli-induced interleukin-8 production.

Application of the FLP/FRT system for conditional gene deletion in yeast Saccharomyces cerevisiae.

PubMed

Park, Yang-Nim; Masison, Daniel; Eisenberg, Evan; Greene, Lois E

2011-09-01

The yeast Saccharomyces cerevisiae has proved to be an excellent model organism to study the function of proteins. One of the many advantages of yeast is the many genetic tools available to manipulate gene expression, but there are still limitations. To complement the many methods used to control gene expression in yeast, we have established a conditional gene deletion system by using the FLP/FRT system on yeast vectors to conditionally delete specific yeast genes. Expression of Flp recombinase, which is under the control of the GAL1 promoter, was induced by galactose, which in turn excised FRT sites flanked genes. The efficacy of this system was examined using the FRT site-flanked genes HSP104, URA3 and GFP. The pre-excision frequency of this system, which might be caused by the basal activity of the GAL1 promoter or by spontaneous recombination between FRT sites, was detected ca. 2% under the non-selecting condition. After inducing expression of Flp recombinase, the deletion efficiency achieved ca. 96% of cells in a population within 9 h. After conditional deletion of the specific gene, protein degradation and cell division then diluted out protein that was expressed from this gene prior to its excision. Most importantly, the specific protein to be deleted could be expressed under its own promoter, so that endogenous levels of protein expression were maintained prior to excision by the Flp recombinase. Therefore, this system provides a useful tool for the conditional deletion of genes in yeast. Published in 2011 by John Wiley & Sons, Ltd.

Yeast cell differentiation: Lessons from pathogenic and non-pathogenic yeasts.

PubMed

Palková, Zdena; Váchová, Libuše

2016-09-01

Yeasts, historically considered to be single-cell organisms, are able to activate different differentiation processes. Individual yeast cells can change their life-styles by processes of phenotypic switching such as the switch from yeast-shaped cells to filamentous cells (pseudohyphae or true hyphae) and the transition among opaque, white and gray cell-types. Yeasts can also create organized multicellular structures such as colonies and biofilms, and the latter are often observed as contaminants on surfaces in industry and medical care and are formed during infections of the human body. Multicellular structures are formed mostly of stationary-phase or slow-growing cells that diversify into specific cell subpopulations that have unique metabolic properties and can fulfill specific tasks. In addition to the development of multiple protective mechanisms, processes of metabolic reprogramming that reflect a changed environment help differentiated individual cells and/or community cell constituents to survive harmful environmental attacks and/or to escape the host immune system. This review aims to provide an overview of differentiation processes so far identified in individual yeast cells as well as in multicellular communities of yeast pathogens of the Candida and Cryptococcus spp. and the Candida albicans close relative, Saccharomyces cerevisiae. Molecular mechanisms and extracellular signals potentially involved in differentiation processes are also briefly mentioned. Copyright © 2016 Elsevier Ltd. All rights reserved.

L-arabinose fermenting yeast

DOEpatents

Zhang, Min; Singh, Arjun; Knoshaug, Eric; Franden, Mary Ann; Jarvis, Eric; Suominen, Pirkko

2010-12-07

An L-arabinose utilizing yeast strain is provided for the production of ethanol by introducing and expressing bacterial araA, araB and araD genes. L-arabinose transporters are also introduced into the yeast to enhance the uptake of arabinose. The yeast carries additional genomic mutations enabling it to consume L-arabinose, even as the only carbon source, and to produce ethanol. Methods of producing ethanol include utilizing these modified yeast strains. ##STR00001##

Insulin-like growth factor I/somatomedin C: a potent inducer of oligodendrocyte development

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

McMorris, F.A.; Smith, T.M.; DeSalvo, S.

1986-02-01

Cell cultures established from cerebrum of 1-day-old rats were used to investigate hormonal regulation of the development of oligodendrocytes, which synthesize myelin in the central nervous system. The number of oligodendrocytes that developed was preferentially increased by insulin, or by insulin-like growth factor I (IGF-I), also known as somatomedin C. High concentrations of insulin were required for substantial induction of oligodendrocyte development, whereas only 3.3 ng of IGF-I per ml was needed for a 2-fold increase in oligodendrocyte numbers. At an IGF-I concentration of 100 ng/ml, oligodendrocyte numbers were increased 6-fold in cultures grown in the presence of 10% fetalmore » bovine serum, or up to 60-fold in cultures maintained in serum-free medium. IGF-I produced less than a 2-fold increase in the number of nonoligodendroglial cells in the same cultures. Type I IGF receptors were identified on oligodendrocytes and on a putative oligodendrocyte precursor cell population identified by using mouse monoclonal antibody A2B5. Radioligand binding assays were done. These results indicate that IGF-I is a potent inducer of oligodendrocyte development and suggest a possible mechanism based on IGF deficiency for the hypomyelination that results from early postnatal malnutrition.« less

Nitrile Metabolizing Yeasts

NASA Astrophysics Data System (ADS)

Bhalla, Tek Chand; Sharma, Monica; Sharma, Nitya Nand

Nitriles and amides are widely distributed in the biotic and abiotic components of our ecosystem. Nitrile form an important group of organic compounds which find their applications in the synthesis of a large number of compounds used as/in pharmaceutical, cosmetics, plastics, dyes, etc>. Nitriles are mainly hydro-lyzed to corresponding amide/acid in organic chemistry. Industrial and agricultural activities have also lead to release of nitriles and amides into the environment and some of them pose threat to human health. Biocatalysis and biotransformations are increasingly replacing chemical routes of synthesis in organic chemistry as a part of ‘green chemistry’. Nitrile metabolizing organisms or enzymes thus has assumed greater significance in all these years to convert nitriles to amides/ acids. The nitrile metabolizing enzymes are widely present in bacteria, fungi and yeasts. Yeasts metabolize nitriles through nitrilase and/or nitrile hydratase and amidase enzymes. Only few yeasts have been reported to possess aldoxime dehydratase. More than sixty nitrile metabolizing yeast strains have been hither to isolated from cyanide treatment bioreactor, fermented foods and soil. Most of the yeasts contain nitrile hydratase-amidase system for metabolizing nitriles. Transformations of nitriles to amides/acids have been carried out with free and immobilized yeast cells. The nitrilases of Torulopsis candida>and Exophiala oligosperma>R1 are enantioselec-tive and regiospecific respectively. Geotrichum>sp. JR1 grows in the presence of 2M acetonitrile and may have potential for application in bioremediation of nitrile contaminated soil/water. The nitrilase of E. oligosperma>R1 being active at low pH (3-6) has shown promise for the hydroxy acids. Immobilized yeast cells hydrolyze some additional nitriles in comparison to free cells. It is expected that more focus in future will be on purification, characterization, cloning, expression and immobilization of nitrile metabolizing

Exploration of potential baker's yeast from sugarcane juice: optimization and evaluation.

PubMed

Mamun-Or-rashid, A N M; Dash, Biplab Kumar; Chowdhury, Md Nurul Abadin; Waheed, Momtaz Fatima; Pramanik, Md Kamruzzaman

2013-07-01

The present study was carried out to explore baker's yeasts strains from sugarcane juice to assess its potential in laboratory scale production of breads. Collected juice samples were processed for isolation and identification of yeast strains based on standard cultural, morphological and biochemical characteristics. Among the six isolated strains, four (designated as S1, S2, S5 and S6) were identified as Saccharomyces cerevisiae and the rests (designated S3 and S4) were as S. rouxii. When assessing their CO2 production rates as a measure of their baking potential, S6 was found to produce maximum amount of gas (226.67 mm3 mL(-1)) in sucrose broth, whereas gas produced by S2, S1 and S5 were relatively insignificant (170, 136.67 and 86.67 mm3 mL(-1), respectively). No strain was found to produce undesirable H2S gas responsible for off-flavor. Besides, effects of different physicochemical parameters (e.g., pH, temperature, substrate concentration, incubation period, agitation etc.) on the production of yeast cell-mass were studied. Yield of cell mass was indirectly measured by spectrophotometric method at 550 nm. All the test isolates were found to produce maximum cell mass at a pH range of 4.0 to 5.0 in 2 to 4% molasses broth at 30 degrees C after 4 days of incubation. In the laboratory scale production of bread using composite flour, Isolate-S6 formed significant characteristic texture. Considering overall characteristics, Isolate- S6 was found to be satisfactorily potent for baking purpose.

Novel Gallate Triphenylphosphonium Derivatives with Potent Antichagasic Activity

PubMed Central

Cortes, Leonel A.; Castro, Lorena; Pesce, Bárbara; Maya, Juan D.; Ferreira, Jorge; Castro-Castillo, Vicente; Parra, Eduardo; Jara, José A.; López-Muñoz, Rodrigo

2015-01-01

Chagas disease is one of the most neglected tropical diseases in the world, affecting nearly 15 million people, primarily in Latin America. Only two drugs are used for the treatment of this disease, nifurtimox and benznidazole. These drugs have limited efficacy and frequently induce adverse effects, limiting their usefulness. Consequently, new drugs must be found. In this study, we demonstrated the in vitro trypanocidal effects of a series of four gallic acid derivatives characterized by a gallate group linked to a triphenylphosphonium (TPP+) moiety (a delocalized cation) via a hydrocarbon chain of 8, 10, 11, or 12 atoms (TPP+-C8, TPP+-C10, TPP+-C11, and TPP+-C12, respectively). We analyzed parasite viability in isolated parasites (by MTT reduction and flow cytometry) and infected mammalian cells using T. cruzi Y strain trypomastigotes. Among the four derivatives, TPP+-C10 and TPP+-C12 were the most potent in both models, with EC50 values (in isolated parasites) of 1.0 ± 0.6 and 1.0 ± 0.7 μM, respectively, and were significantly more potent than nifurtimox (EC50 = 4.1 ± 0.6 μM). At 1 μM, TPP+-C10 and TPP+-C12 induced markers of cell death, such as phosphatidylserine exposure and propidium iodide permeabilization. In addition, at 1 μM, TPP+-C10 and TPP+-C12 significantly decreased the number of intracellular amastigotes (TPP+-C10: 24.3%, TPP+-C12: 19.0% of control measurements, as measured by DAPI staining) and the parasite’s DNA load (C10: 10%, C12: 13% of control measurements, as measured by qPCR). Based on the previous mode of action described for these compounds in cancer cells, we explored their mitochondrial effects in isolated trypomastigotes. TPP+-C10 and TPP+-C12 were the most potent compounds, significantly altering mitochondrial membrane potential at 1 μM (measured by JC-1 fluorescence) and inducing mitochondrial transition pore opening at 5 μM. Taken together, these results indicate that the TPP+-C10 and TPP+-C12 derivatives of gallic acid are

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

Saccharomyces cerevisiae Produces a Yeast Substance that Exhibits Estrogenic Activity in Mammalian Systems

NASA Astrophysics Data System (ADS)

Feldman, David; Stathis, Peter A.; Hirst, Margaret A.; Price Stover, E.; Do, Yung S.; Kurz, Walter

1984-06-01

Partially purified lipid extracts of Saccharomyces cerevisiae contain a substance that displaces tritiated estradiol from rat uterine cytosol estrogen receptors. The yeast product induces estrogenic bioresponses in mammalian systems as measured by induction of progesterone receptors in cultured MCF-7 human breast cancer cells and by a uterotrophic response and progesterone receptor induction after administration to ovariectomized mice. The findings raise the possibility that bakers' yeast may be a source of environmental estrogens.

IL-17 induces osteoclastogenesis from human monocytes alone in the absence of osteoblasts, which is potently inhibited by anti-TNF-alpha antibody: a novel mechanism of osteoclastogenesis by IL-17.

PubMed

Yago, Toru; Nanke, Yuki; Ichikawa, Naomi; Kobashigawa, Tsuyoshi; Mogi, Makio; Kamatani, Naoyuki; Kotake, Shigeru

2009-11-01

IL-17 is a proinflammatory cytokine crucial for osteoclastic bone resorption in the presence of osteoblasts or synoviocytes in rheumatoid arthritis. However, the role of IL-17 in osteoclastogenesis from human monocytes alone remains unclear. Here, we investigated the role of IL-17 in osteoclastogenesis from human monocytes alone and the direct effect of infliximab on the osteoclastogenesis induced by IL-17. Human peripheral blood mononuclear cells (PBMC) were cultured for 3 days with M-CSF. After non-adherent cells were removed, IL-17 was added with either infliximab or osteoprotegerin (OPG). Seven days later, adherent cells were stained for vitronectin receptor. On the other hand, CD11b-positive monocytes purified from PBMC were also cultured and stained as described above. CD11b-positive cells were cultured with TNF-alpha and receptor activator of NF-kappaB ligand (RANKL). In the cultures of both adherent cells and CD11b-positive cells, IL-17 dose-dependently induced osteoclastogenesis in the absence of soluble-RANKL. OPG or infliximab inhibited IL-17-induced osteoclastogenesis. Interestingly, in the culture of CD11b-positive cells, the osteoclastogenesis was more potently inhibited by infliximab than by OPG. TNF-alpha and RANKL synergistically induced osteoclastogenesis. The present study clearly demonstrated the novel mechanism by which IL-17 directly induces osteoclastogenesis from human monocytes alone. In addition, infliximab potently inhibits the osteoclastogenesis directly induced by IL-17. (c) 2009 Wiley-Liss, Inc.

[Relationship between HBeAg from HBsAg positive mothers and regulatory T cells in neonates and its influence on HBV intrauterine transmission].

PubMed

Hao, H Y; Yang, Z Q; Xu, X X; Wang, X F; Wang, B; Shi, X H; Fu, Z D; Wang, B; Wang, S P

2017-10-10

Objective: To explore the relationship between HBeAg in HBsAg positive mothers and CD(4)(+)CD(25)(+) Foxp3 (+)regulatory T cells (Treg) in newborns, as well as how they would influence the increasing risk on HBV intrauterine transmission. Methods: We collected information on general demographic characteristics and delivery on 270 HBsAg positive mothers and their newborns from the Third People's Hospital of Taiyuan. Fluorescence quantitative polymerase chain reaction (FQ-PCR) and chemiluminescence immunoassay (CLIA) were used to detect HBV DNA and HBV serological markers in peripheral blood from both mothers and neonates. The expression of Treg and other immune cells in peripheral blood of neonates were detected with flow cytometry (FCM). Results: Maternal HBeAg positive rates were associated with an increased risk of intrauterine transmission ( OR =4.08, 95 %CI : 1.89-8.82). Rates of Treg in newborns born to HBsAg-positive mothers were higher than that of the negative group ( Z =2.29, P =0.022). Each pair of the subjects was assigned to five different groups according to the HBeAg titers of mothers. Frequencies of both Treg and HBeAg in newborns and HBV DNA in mothers between the above said 5 groups showed similar trends of changing patterns and the differences between groups were statistically significant(χ(2)=18.73, P <0.001; χ(2)=181.60, P <0.001; χ(2)=183.09, P <0.001). Results from partial correlation analysis showed that after adjusting for neonatal HBeAg and maternal HBV DNA, mother's HBeAg titers were positively related to the percentage of Treg in their newborns ( r(s) =0.19, P =0.039). In addition, the frequencies of Treg were negatively correlated with pDC and CD(4)(+) T cell in their newborns ( r(s) =-0.21, P =0.017; r(s) =-0.23, P =0.009). Conclusion: HBeAg from HBsAg positive mothers might have inhibited the function of neonatal DC cells and T cells to reduce the immune response to HBV by up-regulating the proportion of Treg and finally increased

Cadmium is a potent inhibitor of PPM phosphatases and targets the M1 binding site

PubMed Central

Pan, Chang; Liu, Hong-Da; Gong, Zheng; Yu, Xiao; Hou, Xu-Ben; Xie, Di-Dong; Zhu, Xi-Bin; Li, Hao-Wen; Tang, Jun-Yi; Xu, Yun-Fei; Yu, Jia-Qi; Zhang, Lian-Ying; Fang, Hao; Xiao, Kun-Hong; Chen, Yu-Guo; Wang, Jiang-Yun; Pang, Qi; Chen, Wei; Sun, Jin-Peng

2013-01-01

The heavy metal cadmium is a non-degradable pollutant. By screening the effects of a panel of metal ions on the phosphatase activity, we unexpectedly identified cadmium as a potent inhibitor of PPM1A and PPM1G. In contrast, low micromolar concentrations of cadmium did not inhibit PP1 or tyrosine phosphatases. Kinetic studies revealed that cadmium inhibits PPM phosphatases through the M1 metal ion binding site. In particular, the negative charged D441 in PPM1G specific recognized cadmium. Our results suggest that cadmium is likely a potent inhibitor of most PPM family members except for PHLPPs. Furthermore, we demonstrated that cadmium inhibits PPM1A-regulated MAPK signaling and PPM1G-regulated AKT signaling potently in vivo. Cadmium reversed PPM1A-induced cell cycle arrest and cadmium insensitive PPM1A mutant rescued cadmium induced cell death. Taken together, these findings provide a better understanding of the effects of the toxicity of cadmium in the contexts of human physiology and pathology. PMID:23903585

The essence of yeast quiescence.

PubMed

De Virgilio, Claudio

2012-03-01

Like all microorganisms, yeast cells spend most of their natural lifetime in a reversible, quiescent state that is primarily induced by limitation for essential nutrients. Substantial progress has been made in defining the features of quiescent cells and the nutrient-signaling pathways that shape these features. A view that emerges from the wealth of new data is that yeast cells dynamically configure the quiescent state in response to nutritional challenges by using a set of key nutrient-signaling pathways, which (1) regulate pathway-specific effectors, (2) converge on a few regulatory nodes that bundle multiple inputs to communicate unified, graded responses, and (3) mutually modulate their competences to transmit signals. Here, I present an overview of our current understanding of the architecture of these pathways, focusing on how the corresponding core signaling protein kinases (i.e. PKA, TORC1, Snf1, and Pho85) are wired to ensure an adequate response to nutrient starvation, which enables cells to tide over decades, if not centuries, of famine. © 2011 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

New yeasts-new brews: modern approaches to brewing yeast design and development.

PubMed

Gibson, B; Geertman, J-M A; Hittinger, C T; Krogerus, K; Libkind, D; Louis, E J; Magalhães, F; Sampaio, J P

2017-06-01

The brewing industry is experiencing a period of change and experimentation largely driven by customer demand for product diversity. This has coincided with a greater appreciation of the role of yeast in determining the character of beer and the widespread availability of powerful tools for yeast research. Genome analysis in particular has helped clarify the processes leading to domestication of brewing yeast and has identified domestication signatures that may be exploited for further yeast development. The functional properties of non-conventional yeast (both Saccharomyces and non-Saccharomyces) are being assessed with a view to creating beers with new flavours as well as producing flavoursome non-alcoholic beers. The discovery of the psychrotolerant S. eubayanus has stimulated research on de novo S. cerevisiae × S. eubayanus hybrids for low-temperature lager brewing and has led to renewed interest in the functional importance of hybrid organisms and the mechanisms that determine hybrid genome function and stability. The greater diversity of yeast that can be applied in brewing, along with an improved understanding of yeasts' evolutionary history and biology, is expected to have a significant and direct impact on the brewing industry, with potential for improved brewing efficiency, product diversity and, above all, customer satisfaction. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

DNA vaccines targeting the encoded antigens to dendritic cells induce potent antitumor immunity in mice.

PubMed

Cao, Jun; Jin, Yiqi; Li, Wei; Zhang, Bin; He, Yang; Liu, Hongqiang; Xia, Ning; Wei, Huafeng; Yan, Jian

2013-08-14

Although DNA vaccine holds a great potential for cancer immunotherapy, effective long-lasting antitumoral immunity sufficient to induce durable responses in cancer patients remains to be achieved. Considering the pivotal role of dendritic cells (DC) in the antigen processing and presentation, we prepared DC-targeting DNA vaccines by fusing tumor-associated antigen HER2/neu ectodomain to single chain antibody fragment (scFv) from NLDC-145 antibody specific for DC-restricted surface molecule DEC-205 (scFvNLDC-145), and explored its antitumoral efficacy and underlying mechanisms in mouse breast cancer models. In vivo targeting assay demonstrated that scFvNLDC-145 specifically delivered DNA vaccine-encoded antigen to DC. Compared with untargeted HER2/neu DNA vaccines, vaccination with scFvNLDC-145-HER2/neu markedly promoted the HER2/neu-specific cellular and humoral immune responses with long-lasting immune memory, resulting in effective protection against challenge of HER2/neu-positive D2F2/E2 breast tumor while ineffective in parental HER2/neu-negative D2F2 breast tumor. More importantly, in combination with temporary depletion of regulatory T cells (Treg) by low-dose cyclophosphamide, vaccination with scFvNLDC-145-HER2/neu induced the regression of established D2F2/E2 breast tumor and significantly retarded the development of spontaneous mammary carcinomas in transgenic BALB-neuT mice. Our findings demonstrate that DC-targeted DNA vaccines for in vivo direct delivery of tumor antigens to DC could induce potent antigen-specific cellular and humoral immune responses and, if additional combination with systemic Treg depletion, was able to elicit an impressively therapeutic antitumoral activity, providing a rationale for further development of this approach for cancer treatment.

S-Adenosyl-L-methionine protects the probiotic yeast, Saccharomyces boulardii, from acid-induced cell death.

PubMed

Cascio, Vincent; Gittings, Daniel; Merloni, Kristen; Hurton, Matthew; Laprade, David; Austriaco, Nicanor

2013-02-13

Saccharomyces boulardii is a probiotic yeast routinely used to prevent and to treat gastrointestinal disorders, including the antibiotic-associated diarrhea caused by Clostridium difficile infections. However, only 1-3% of the yeast administered orally is recovered alive in the feces suggesting that this yeast is unable to survive the acidic environment of the gastrointestinal tract. We provide evidence that suggests that S. boulardii undergoes programmed cell death (PCD) in acidic environments, which is accompanied by the generation of reactive oxygen species and the appearance of caspase-like activity. To better understand the mechanism of cell death at the molecular level, we generated microarray gene expression profiles of S. boulardii cells cultured in an acidic environment. Significantly, functional annotation revealed that the up-regulated genes were significantly over-represented in cell death pathways Finally, we show that S-adenosyl-L-methionine (AdoMet), a commercially available, FDA-approved dietary supplement, enhances the viability of S. boulardii in acidic environments, most likely by preventing programmed cell death. In toto, given the observation that many of the proven health benefits of S. boulardii are dependent on cell viability, our data suggests that taking S. boulardii and AdoMet together may be a more effective treatment for gastrointestinal disorders than taking the probiotic yeast alone.

Isolated yeast promoter sequence and a method of regulated heterologous expression

DOEpatents

Gao, Johnway; Skeen, Rodney S.; Hooker, Brian S.; Anderson, Daniel B.

2005-05-31

The present invention provides the promoter clone discovery of a glucoamylase gene of a starch utilizing yeast strain Schwanniomyces castellii. The isolated glucoamylase promoter is an inducible promoter, which can regulate strong gene expression in starch culture medium.

Glycation inhibitors extend yeast chronological lifespan by reducing advanced glycation end products and by back regulation of proteins involved in mitochondrial respiration.

PubMed

Kazi, Rubina S; Banarjee, Reema M; Deshmukh, Arati B; Patil, Gouri V; Jagadeeshaprasad, Mashanipalya G; Kulkarni, Mahesh J

2017-03-06

Advanced Glycation End products (AGEs) are implicated in aging process. Thus, reducing AGEs by using glycation inhibitors may help in attenuating the aging process. In this study using Saccharomyces cerevisiae yeast system, we show that Aminoguanidine (AMG), a well-known glycation inhibitor, decreases the AGE modification of proteins in non-calorie restriction (NR) (2% glucose) and extends chronological lifespan (CLS) similar to that of calorie restriction (CR) condition (0.5% glucose). Proteomic analysis revealed that AMG back regulates the expression of differentially expressed proteins especially those involved in mitochondrial respiration in NR condition, suggesting that it switches metabolism from fermentation to respiration, mimicking CR. AMG induced back regulation of differentially expressed proteins could be possibly due to its chemical effect or indirectly by glycation inhibition. To delineate this, Metformin (MET), a structural analog of AMG and a mild glycation inhibitor and Hydralazine (HYD), another potent glycation inhibitor but not structural analog of AMG were used. HYD was more effective than MET in mimicking AMG suggesting that glycation inhibition was responsible for restoration of differentially expressed proteins. Thus glycation inhibitors particularly AMG, HYD and MET extend yeast CLS by reducing AGEs, modulating the expression of proteins involved in mitochondrial respiration and possibly by scavenging glucose. This study reports the role of glycation in aging process. In the non-caloric restriction condition, carbohydrates such as glucose promote protein glycation and reduce CLS. While, the inhibitors of glycation such as AMG, HYD, MET mimic the caloric restriction condition by back regulating deregulated proteins involved in mitochondrial respiration which could facilitate shift of metabolism from fermentation to respiration and extend yeast CLS. These findings suggest that glycation inhibitors can be potential molecules that can be used

METHYLATION INACTIVATES PENTAVALENT ARSENIC SPECIES BUT ACTIVATES TRIVALENT ARSENIC SPECIES TO POTENT GENOTOXICANTS

EPA Science Inventory

Methylation Inactivates Pentavalent Arsenic Species but Activates Trivalent Arsenic Species to Potent Genotoxicants

The sensitivity ofhumans to arsenic-induced cancer is thought to be related in part to the limited ability of humans to detoxify arsenic. Recently, methyl- ...

Lemon grass (Cymbopogon citratus) essential oil as a potent anti-inflammatory and antifungal drugs

PubMed Central

Boukhatem, Mohamed Nadjib; Ferhat, Mohamed Amine; Kameli, Abdelkrim; Saidi, Fairouz; Kebir, Hadjer Tchoketch

2014-01-01

Background Volatile oils obtained from lemon grass [Cymbopogon citratus (DC.) Stapf, Poaceae family] are used in traditional medicine as remedies for the treatment of various diseases. Aims In the present study, lemon grass essential oil (LGEO) was evaluated for its in vivo topical and oral anti-inflammatory effects, and for its in vitro antifungal activity using both liquid and vapor phases. Methods The chemical profile of LGEO as determined by gas chromatography–mass spectrometry analysis revealed two major components: geranial (42.2%), and neral (31.5%). The antifungal activity of LGEO was evaluated against several pathogenic yeasts and filamentous fungi using disc diffusion and vapor diffusion methods. Results LGEO exhibited promising antifungal effect against Candida albicans, C. tropicalis, and Aspergillus niger, with different inhibition zone diameters (IZDs) (35–90 mm). IZD increased with increasing oil volume. Significantly, higher anti-Candida activity was observed in the vapor phase. For the evaluation of the anti-inflammatory effect, LGEO (10 mg/kg, administered orally) significantly reduced carrageenan-induced paw edema with a similar effect to that observed for oral diclofenac (50 mg/kg), which was used as the positive control. Oral administration of LGEO showed dose-dependent anti-inflammatory activity. In addition, topical application of LGEO in vivo resulted in a potent anti-inflammatory effect, as demonstrated by using the mouse model of croton oil-induced ear edema. To our knowledge, this is the first such report to be published. The topical application of LGEO at doses of 5 and 10 µL/ear significantly reduced acute ear edema induced by croton oil in 62.5 and 75% of the mice, respectively. In addition, histological analysis clearly confirmed that LGEO inhibits the skin inflammatory response in animal models. Conclusion Results of the present study indicate that LGEO has a noteworthy potential for the development of drugs for the treatment of

Lemon grass (Cymbopogon citratus) essential oil as a potent anti-inflammatory and antifungal drugs.

PubMed

Boukhatem, Mohamed Nadjib; Ferhat, Mohamed Amine; Kameli, Abdelkrim; Saidi, Fairouz; Kebir, Hadjer Tchoketch

2014-01-01

Background Volatile oils obtained from lemon grass [Cymbopogon citratus (DC.) Stapf, Poaceae family] are used in traditional medicine as remedies for the treatment of various diseases. Aims In the present study, lemon grass essential oil (LGEO) was evaluated for its in vivo topical and oral anti-inflammatory effects, and for its in vitro antifungal activity using both liquid and vapor phases. Methods The chemical profile of LGEO as determined by gas chromatography-mass spectrometry analysis revealed two major components: geranial (42.2%), and neral (31.5%). The antifungal activity of LGEO was evaluated against several pathogenic yeasts and filamentous fungi using disc diffusion and vapor diffusion methods. Results LGEO exhibited promising antifungal effect against Candida albicans, C.tropicalis, and Aspergillus niger, with different inhibition zone diameters (IZDs) (35-90 mm). IZD increased with increasing oil volume. Significantly, higher anti-Candida activity was observed in the vapor phase. For the evaluation of the anti-inflammatory effect, LGEO (10 mg/kg, administered orally) significantly reduced carrageenan-induced paw edema with a similar effect to that observed for oral diclofenac (50 mg/kg), which was used as the positive control. Oral administration of LGEO showed dose-dependent anti-inflammatory activity. In addition, topical application of LGEO in vivo resulted in a potent anti-inflammatory effect, as demonstrated by using the mouse model of croton oil-induced ear edema. To our knowledge, this is the first such report to be published. The topical application of LGEO at doses of 5 and 10 µL/ear significantly reduced acute ear edema induced by croton oil in 62.5 and 75% of the mice, respectively. In addition, histological analysis clearly confirmed that LGEO inhibits the skin inflammatory response in animal models. Conclusion Results of the present study indicate that LGEO has a noteworthy potential for the development of drugs for the treatment of

Lemon grass (Cymbopogon citratus) essential oil as a potent anti-inflammatory and antifungal drugs.

PubMed

Boukhatem, Mohamed Nadjib; Ferhat, Mohamed Amine; Kameli, Abdelkrim; Saidi, Fairouz; Kebir, Hadjer Tchoketch

2014-01-01

Volatile oils obtained from lemon grass [Cymbopogon citratus (DC.) Stapf, Poaceae family] are used in traditional medicine as remedies for the treatment of various diseases. In the present study, lemon grass essential oil (LGEO) was evaluated for its in vivo topical and oral anti-inflammatory effects, and for its in vitro antifungal activity using both liquid and vapor phases. The chemical profile of LGEO as determined by gas chromatography-mass spectrometry analysis revealed two major components: geranial (42.2%), and neral (31.5%). The antifungal activity of LGEO was evaluated against several pathogenic yeasts and filamentous fungi using disc diffusion and vapor diffusion methods. LGEO exhibited promising antifungal effect against Candida albicans, C. tropicalis, and Aspergillus niger, with different inhibition zone diameters (IZDs) (35-90 mm). IZD increased with increasing oil volume. Significantly, higher anti-Candida activity was observed in the vapor phase. For the evaluation of the anti-inflammatory effect, LGEO (10 mg/kg, administered orally) significantly reduced carrageenan-induced paw edema with a similar effect to that observed for oral diclofenac (50 mg/kg), which was used as the positive control. Oral administration of LGEO showed dose-dependent anti-inflammatory activity. In addition, topical application of LGEO in vivo resulted in a potent anti-inflammatory effect, as demonstrated by using the mouse model of croton oil-induced ear edema. To our knowledge, this is the first such report to be published. The topical application of LGEO at doses of 5 and 10 µL/ear significantly reduced acute ear edema induced by croton oil in 62.5 and 75% of the mice, respectively. In addition, histological analysis clearly confirmed that LGEO inhibits the skin inflammatory response in animal models. RESULTS of the present study indicate that LGEO has a noteworthy potential for the development of drugs for the treatment of fungal infections and skin inflammation that

A survey of liver pathology in needle biopsies from HBsAg and anti-HBe positive individuals.

PubMed

ter Borg, F; ten Kate, F J; Cuypers, H T; Leentvaar-Kuijpers, A; Oosting, J; Wertheim-van Dillen, P M; Honkoop, P; Rasch, M C; de Man, R A; van Hattum, J; Chamuleau, R A; Tytgat, G N; Jones, E A

2000-07-01

To use laboratory data and liver biopsies, prospectively obtained from hepatitis B surface antigen (HBsAg) and anti hepatitis B e antigen (anti-HBe) positive patients, for the assessment of: (1) the relation between biopsy length/number of portal tracts and sampling error; (2) the relation between the severity of piecemeal necrosis and the new grading terminology (minimal, mild, moderate, and severe chronic hepatitis); and (3) liver pathology, which has not been studied in patients with this specific serological profile. The study group (n = 174) included 104 patients with normal aminotransferase concentrations and no cases with clinically apparent cirrhosis. The specimen length and number of portal tracts were measured at light microscopy examination. Sampling error analysis was related to the discrepancies between aminotransferase concentrations versus histological grade. Detailed histological scorings were undertaken by the reference pathologist and compared with laboratory and hepatitis B virus (HBV) DNA precore sequence data. Sampling error seemed to be a constant feature, even for biopsies > or = 20 mm, but increased dramatically in biopsies < 5 mm long and/or containing less than four portal tracts. Between 25% and 30% of biopsies, graded as "mild" or "moderate" activity showed features of moderate and severe piecemeal necrosis, respectively. Ten per cent of the patients with normal aminotransferase values had stage III-IV hepatic fibrosis, and 20% had piecemeal necrosis. Only cytoplasmic, not nuclear, core antigen expression was a strong predictor of high hepatitis B viraemia. There was no association between precore stop codon mutations, grade/stage of liver disease, and hepatitis B core antigen (HBcAg) expression. The specimen available for light microscopical examination should be > 5 mm long and should contain more than four portal tracts. In addition, the new grading terminology might give the clinician an inappropriately mild impression of the

Vaginal yeast infection

MedlinePlus

Yeast infection - vagina; Vaginal candidiasis; Monilial vaginitis ... Most women have a vaginal yeast infection at some time. Candida albicans is a common type of fungus. It is often found in small amounts in the ...

The proline metabolism intermediate Δ1-pyrroline-5-carboxylate directly inhibits the mitochondrial respiration in budding yeast.

PubMed

Nishimura, Akira; Nasuno, Ryo; Takagi, Hiroshi

2012-07-30

The proline metabolism intermediate Δ(1)-pyrroline-5-carboxylate (P5C) induces cell death in animals, plants and yeasts. To elucidate how P5C triggers cell death, we analyzed P5C metabolism, mitochondrial respiration and superoxide anion generation in the yeast Saccharomyces cerevisiae. Gene disruption analysis revealed that P5C-mediated cell death was not due to P5C metabolism. Interestingly, deficiency in mitochondrial respiration suppressed the sensitivity of yeast cells to P5C. In addition, we found that P5C inhibits the mitochondrial respiration and induces a burst of superoxide anions from the mitochondria. We propose that P5C regulates cell death via the inhibition of mitochondrial respiration. Copyright © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

DNA strand-exchange patterns associated with double-strand break-induced and spontaneous mitotic crossovers in Saccharomyces cerevisiae

PubMed Central

2018-01-01

Mitotic recombination can result in loss of heterozygosity and chromosomal rearrangements that shape genome structure and initiate human disease. Engineered double-strand breaks (DSBs) are a potent initiator of recombination, but whether spontaneous events initiate with the breakage of one or both DNA strands remains unclear. In the current study, a crossover (CO)-specific assay was used to compare heteroduplex DNA (hetDNA) profiles, which reflect strand exchange intermediates, associated with DSB-induced versus spontaneous events in yeast. Most DSB-induced CO products had the two-sided hetDNA predicted by the canonical DSB repair model, with a switch in hetDNA position from one product to the other at the position of the break. Approximately 40% of COs, however, had hetDNA on only one side of the initiating break. This anomaly can be explained by a modified model in which there is frequent processing of an early invasion (D-loop) intermediate prior to extension of the invading end. Finally, hetDNA tracts exhibited complexities consistent with frequent expansion of the DSB into a gap, migration of strand-exchange junctions, and template switching during gap-filling reactions. hetDNA patterns in spontaneous COs isolated in either a wild-type background or in a background with elevated levels of reactive oxygen species (tsa1Δ mutant) were similar to those associated with the DSB-induced events, suggesting that DSBs are the major instigator of spontaneous mitotic recombination in yeast. PMID:29579095

Identification and characterisation of xylanolytic yeasts isolated from decaying wood and sugarcane bagasse in Brazil.

PubMed

Lara, Carla A; Santos, Renata O; Cadete, Raquel M; Ferreira, Carla; Marques, Susana; Gírio, Francisco; Oliveira, Evelyn S; Rosa, Carlos A; Fonseca, César

2014-06-01

In this study, yeasts associated with lignocellulosic materials in Brazil, including decaying wood and sugarcane bagasse, were isolated, and their ability to produce xylanolytic enzymes was investigated. A total of 358 yeast isolates were obtained, with 198 strains isolated from decaying wood and 160 strains isolated from decaying sugarcane bagasse samples. Seventy-five isolates possessed xylanase activity in solid medium and were identified as belonging to nine species: Candida intermedia, C. tropicalis, Meyerozyma guilliermondii, Scheffersomyces shehatae, Sugiyamaella smithiae, Cryptococcus diffluens, Cr. heveanensis, Cr. laurentii and Trichosporon mycotoxinivorans. Twenty-one isolates were further screened for total xylanase activity in liquid medium with xylan, and five xylanolytic yeasts were selected for further characterization, which included quantitative analysis of growth in xylan and xylose and xylanase and β-D-xylosidase activities. The yeasts showing the highest growth rate and cell density in xylan, Cr. laurentii UFMG-HB-48, Su. smithiae UFMG-HM-80.1 and Sc. shehatae UFMG-HM-9.1a, were, simultaneously, those exhibiting higher xylanase activity. Xylan induced the highest level of (extracellular) xylanase activity in Cr. laurentii UFMG-HB-48 and the highest level of (intracellular, extracellular and membrane-associated) β-D-xylosidase activity in Su. smithiae UFMG-HM-80.1. Also, significant β-D-xylosidase levels were detected in xylan-induced cultures of Cr. laurentii UFMG-HB-48 and Sc. shehatae UFMG-HM-9.1a, mainly in extracellular and intracellular spaces, respectively. Under xylose induction, Cr. laurentii UFMG-HB-48 showed the highest intracellular β-D-xylosidase activity among all the yeast tested. C. tropicalis UFMG-HB 93a showed its higher (intracellular) β-D-xylosidase activity under xylose induction and higher at 30 °C than at 50 °C. This study revealed different xylanolytic abilities and strategies in yeasts to metabolise xylan and

Extensive screening for herbal extracts with potent antioxidant properties

PubMed Central

Niwano, Yoshimi; Saito, Keita; Yoshizaki, Fumihiko; Kohno, Masahiro; Ozawa, Toshihiko

2011-01-01

This paper summarizes our research for herbal extracts with potent antioxidant activity obtained from a large scale screening based on superoxide radical (O2•−) scavenging activity followed by characterization of antioxidant properties. Firstly, scavenging activity against O2•− was extensively screened from ethanol extracts of approximately 1000 kinds of herbs by applying an electron spin resonance (ESR)-spin trapping method, and we chose four edible herbal extracts with prominently potent ability to scavenge O2•−. They are the extracts from Punica granatum (Peel), Syzygium aromaticum (Bud), Mangifera indica (Kernel), and Phyllanthus emblica (Fruit). These extracts were further examined to determine if they also scavenge hydroxyl radical (•OH), by applying the ESR spin-trapping method, and if they have heat resistance as a desirable characteristic feature. Experiments with the Fenton reaction and photolysis of H2O2 induced by UV irradiation demonstrated that all four extracts have potent ability to directly scavenge •OH. Furthermore, the scavenging activities against O2•− and •OH of the extracts of P. granatum (peel), M. indica (kernel) and P. emblica (fruit) proved to be heat-resistant. The results of the review might give useful information when choosing a potent antioxidant as a foodstuff. For instance, the four herbal extracts chosen from extensive screening possess desirable antioxidant properties. In particular, the extracts of the aforementioned three herbs are expected to be suitable for food processing in which thermal devices are used, because of their heat resistance. PMID:21297917

Schizosaccharomyces japonicus: the fission yeast is a fusion of yeast and hyphae.

PubMed

Niki, Hironori

2014-03-01

The clade of Schizosaccharomyces includes 4 species: S. pombe, S. octosporus, S. cryophilus, and S. japonicus. Although all 4 species exhibit unicellular growth with a binary fission mode of cell division, S. japonicus alone is dimorphic yeast, which can transit from unicellular yeast to long filamentous hyphae. Recently it was found that the hyphal cells response to light and then synchronously activate cytokinesis of hyphae. In addition to hyphal growth, S. japonicas has many properties that aren't shared with other fission yeast. Mitosis of S. japonicas is referred to as semi-open mitosis because dynamics of nuclear membrane is an intermediate mode between open mitosis and closed mitosis. Novel genetic tools and the whole genomic sequencing of S. japonicas now provide us with an opportunity for revealing unique characters of the dimorphic yeast. © 2013 The Author. Yeast Published by John Wiley & Sons Ltd.

Simvastatin Potently Induces Calcium-dependent Apoptosis of Human Leiomyoma Cells*

PubMed Central

Borahay, Mostafa A.; Kilic, Gokhan S.; Yallampalli, Chandrasekha; Snyder, Russell R.; Hankins, Gary D. V.; Al-Hendy, Ayman; Boehning, Darren

2014-01-01

Statins are drugs commonly used for the treatment of high plasma cholesterol levels. Beyond these well known lipid-lowering properties, they possess broad-reaching effects in vivo, including antitumor effects. Statins inhibit the growth of multiple tumors. However, the mechanisms remain incompletely understood. Here we show that simvastatin inhibits the proliferation of human leiomyoma cells. This was associated with decreased mitogen-activated protein kinase signaling and multiple changes in cell cycle progression. Simvastatin potently stimulated leiomyoma cell apoptosis in a manner mechanistically dependent upon apoptotic calcium release from voltage-gated calcium channels. Therefore, simvastatin possesses antitumor effects that are dependent upon the apoptotic calcium release machinery. PMID:25359773

Marine yeast isolation and industrial application.

PubMed

Zaky, Abdelrahman Saleh; Tucker, Gregory A; Daw, Zakaria Yehia; Du, Chenyu

2014-09-01

Over the last century, terrestrial yeasts have been widely used in various industries, such as baking, brewing, wine, bioethanol and pharmaceutical protein production. However, only little attention has been given to marine yeasts. Recent research showed that marine yeasts have several unique and promising features over the terrestrial yeasts, for example higher osmosis tolerance, higher special chemical productivity and production of industrial enzymes. These indicate that marine yeasts have great potential to be applied in various industries. This review gathers the most recent techniques used for marine yeast isolation as well as the latest applications of marine yeast in bioethanol, pharmaceutical and enzyme production fields. © 2014 The Authors FEMS Yeast Research published by John Wiley & Sons Ltd on behalf of Federation of European Microbiological Societies.

Yeast ecology of Kombucha fermentation.

PubMed

Teoh, Ai Leng; Heard, Gillian; Cox, Julian

2004-09-01

Kombucha is a traditional fermentation of sweetened tea, involving a symbiosis of yeast species and acetic acid bacteria. Despite reports of different yeast species being associated with the fermentation, little is known of the quantitative ecology of yeasts in Kombucha. Using oxytetracycline-supplemented malt extract agar, yeasts were isolated from four commercially available Kombucha products and identified using conventional biochemical and physiological tests. During the fermentation of each of the four products, yeasts were enumerated from both the cellulosic pellicle and liquor of the Kombucha. The number and diversity of species varied between products, but included Brettanomyces bruxellensis, Candida stellata, Schizosaccharomyces pombe, Torulaspora delbrueckii and Zygosaccharomyces bailii. While these yeast species are known to occur in Kombucha, the enumeration of each species present throughout fermentation of each of the four Kombucha cultures demonstrated for the first time the dynamic nature of the yeast ecology. Kombucha fermentation is, in general, initiated by osmotolerant species, succeeded and ultimately dominated by acid-tolerant species.

Antigenic characterisation of yeast-expressed lyssavirus nucleoproteins.

PubMed

Kucinskaite, Indre; Juozapaitis, Mindaugas; Serva, Andrius; Zvirbliene, Aurelija; Johnson, Nicholas; Staniulis, Juozas; Fooks, Anthony R; Müller, Thomas; Sasnauskas, Kestutis; Ulrich, Rainer G

2007-12-01

In Europe, three genotypes of the genus Lyssavirus, family Rhabdoviridae, are present, classical rabies virus (RABV, genotype 1), European bat lyssavirus type 1 (EBLV-1, genotype 5) and European bat lyssavirus type 2 (EBLV-2, genotype 6). The entire authentic nucleoprotein (N protein) encoding sequences of RABV (challenge virus standard, CVS, strain), EBLV-1 and EBLV-2 were expressed in yeast Saccharomyces cerevisiae at high level. Purification of recombinant N proteins by caesium chloride gradient centrifugation resulted in yields between 14-17, 25-29 and 18-20 mg/l of induced yeast culture for RABV-CVS, EBLV-1 and EBLV-2, respectively. The purified N proteins were evaluated by negative staining electron microscopy, which revealed the formation of nucleocapsid-like structures. The antigenic conformation of the N proteins was investigated for their reactivity with monoclonal antibodies (mAbs) directed against different lyssaviruses. The reactivity pattern of each mAb was virtually identical between immunofluorescence assay with virus-infected cells, and ELISA and dot blot assay using the corresponding recombinant N proteins. These observations lead us to conclude that yeast-expressed lyssavirus N proteins share antigenic properties with naturally expressed virus protein. These recombinant proteins have the potential for use as components of serological assays for lyssaviruses.

Marine yeast isolation and industrial application

PubMed Central

Zaky, Abdelrahman Saleh; Tucker, Gregory A; Daw, Zakaria Yehia; Du, Chenyu

2014-01-01

Over the last century, terrestrial yeasts have been widely used in various industries, such as baking, brewing, wine, bioethanol and pharmaceutical protein production. However, only little attention has been given to marine yeasts. Recent research showed that marine yeasts have several unique and promising features over the terrestrial yeasts, for example higher osmosis tolerance, higher special chemical productivity and production of industrial enzymes. These indicate that marine yeasts have great potential to be applied in various industries. This review gathers the most recent techniques used for marine yeast isolation as well as the latest applications of marine yeast in bioethanol, pharmaceutical and enzyme production fields. PMID:24738708

S-Adenosyl-L-Methionine protects the probiotic yeast, Saccharomyces boulardii, from acid-induced cell death

PubMed Central

2013-01-01

Background Saccharomyces boulardii is a probiotic yeast routinely used to prevent and to treat gastrointestinal disorders, including the antibiotic-associated diarrhea caused by Clostridium difficile infections. However, only 1-3% of the yeast administered orally is recovered alive in the feces suggesting that this yeast is unable to survive the acidic environment of the gastrointestinal tract. Results We provide evidence that suggests that S. boulardii undergoes programmed cell death (PCD) in acidic environments, which is accompanied by the generation of reactive oxygen species and the appearance of caspase-like activity. To better understand the mechanism of cell death at the molecular level, we generated microarray gene expression profiles of S. boulardii cells cultured in an acidic environment. Significantly, functional annotation revealed that the up-regulated genes were significantly over-represented in cell death pathways Finally, we show that S-adenosyl-L-methionine (AdoMet), a commercially available, FDA-approved dietary supplement, enhances the viability of S. boulardii in acidic environments, most likely by preventing programmed cell death. Conclusions In toto, given the observation that many of the proven health benefits of S. boulardii are dependent on cell viability, our data suggests that taking S. boulardii and AdoMet together may be a more effective treatment for gastrointestinal disorders than taking the probiotic yeast alone. PMID:23402325

Effects of intracellular iron overload on cell death and identification of potent cell death inhibitors.

PubMed

Fang, Shenglin; Yu, Xiaonan; Ding, Haoxuan; Han, Jianan; Feng, Jie

2018-06-11

Iron overload causes many diseases, while the underlying etiologies of these diseases are unclear. Cell death processes including apoptosis, necroptosis, cyclophilin D-(CypD)-dependent necrosis and a recently described additional form of regulated cell death called ferroptosis, are dependent on iron or iron-dependent reactive oxygen species (ROS). However, whether the accumulation of intracellular iron itself induces ferroptosis or other forms of cell death is largely elusive. In present study, we study the role of intracellular iron overload itself-induced cell death mechanisms by using ferric ammonium citrate (FAC) and a membrane-permeable Ferric 8-hydroxyquinoline complex (Fe-8HQ) respectively. We show that FAC-induced intracellular iron overload causes ferroptosis. We also identify 3-phosphoinositide-dependent kinase 1 (PDK1) inhibitor GSK2334470 as a potent ferroptosis inhibitor. Whereas, Fe-8HQ-induced intracellular iron overload causes unregulated necrosis, but partially activates PARP-1 dependent parthanatos. Interestingly, we identify many phenolic compounds as potent inhibitors of Fe-8HQ-induced cell death. In conclusion, intracellular iron overload-induced cell death form might be dependent on the intracellular iron accumulation rate, newly identified cell death inhibitors in our study that target ferroptosis and unregulated oxidative cell death represent potential therapeutic strategies against iron overload related diseases. Copyright © 2018 Elsevier Inc. All rights reserved.

In vitro assay using engineered yeast vacuoles for neuronal SNARE-mediated membrane fusion

PubMed Central

Ko, Young-Joon; Lee, Miriam; Kang, KyeongJin; Song, Woo Keun; Jun, Youngsoo

2014-01-01

Intracellular membrane fusion requires not only SNARE proteins but also other regulatory proteins such as the Rab and Sec1/Munc18 (SM) family proteins. Although neuronal SNARE proteins alone can drive the fusion between synthetic liposomes, it remains unclear whether they are also sufficient to induce the fusion of biological membranes. Here, through the use of engineered yeast vacuoles bearing neuronal SNARE proteins, we show that neuronal SNAREs can induce membrane fusion between yeast vacuoles and that this fusion does not require the function of the Rab protein Ypt7p or the SM family protein Vps33p, both of which are essential for normal yeast vacuole fusion. Although excess vacuolar SNARE proteins were also shown to mediate Rab-bypass fusion, this fusion required homotypic fusion and vacuole protein sorting complex, which bears Vps33p and was accompanied by extensive membrane lysis. We also show that this neuronal SNARE-driven vacuole fusion can be stimulated by the neuronal SM protein Munc18 and blocked by botulinum neurotoxin serotype E, a well-known inhibitor of synaptic vesicle fusion. Taken together, our results suggest that neuronal SNARE proteins are sufficient to induce biological membrane fusion, and that this new assay can be used as a simple and complementary method for investigating synaptic vesicle fusion mechanisms. PMID:24821814

Wine yeasts for the future.

PubMed

Fleet, Graham H

2008-11-01

International competition within the wine market, consumer demands for newer styles of wines and increasing concerns about the environmental sustainability of wine production are providing new challenges for innovation in wine fermentation. Within the total production chain, the alcoholic fermentation of grape juice by yeasts is a key process where winemakers can creatively engineer wine character and value through better yeast management and, thereby, strategically tailor wines to a changing market. This review considers the importance of yeast ecology and yeast metabolic reactions in determining wine quality, and then discusses new directions for exploiting yeasts in wine fermentation. It covers criteria for selecting and developing new commercial strains, the possibilities of using yeasts other than those in the genus of Saccharomyces, the prospects for mixed culture fermentations and explores the possibilities for high cell density, continuous fermentations.

Global analysis of the yeast lipidome by quantitative shotgun mass spectrometry.

PubMed

Ejsing, Christer S; Sampaio, Julio L; Surendranath, Vineeth; Duchoslav, Eva; Ekroos, Kim; Klemm, Robin W; Simons, Kai; Shevchenko, Andrej

2009-02-17

Although the transcriptome, proteome, and interactome of several eukaryotic model organisms have been described in detail, lipidomes remain relatively uncharacterized. Using Saccharomyces cerevisiae as an example, we demonstrate that automated shotgun lipidomics analysis enabled lipidome-wide absolute quantification of individual molecular lipid species by streamlined processing of a single sample of only 2 million yeast cells. By comparative lipidomics, we achieved the absolute quantification of 250 molecular lipid species covering 21 major lipid classes. This analysis provided approximately 95% coverage of the yeast lipidome achieved with 125-fold improvement in sensitivity compared with previous approaches. Comparative lipidomics demonstrated that growth temperature and defects in lipid biosynthesis induce ripple effects throughout the molecular composition of the yeast lipidome. This work serves as a resource for molecular characterization of eukaryotic lipidomes, and establishes shotgun lipidomics as a powerful platform for complementing biochemical studies and other systems-level approaches.

The flavoprotein Tah18-dependent NO synthesis confers high-temperature stress tolerance on yeast cells

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

Nishimura, Akira; Kawahara, Nobuhiro; Takagi, Hiroshi, E-mail: hiro@bs.naist.jp

Highlights: Black-Right-Pointing-Pointer NO is produced from L-arginine in response to elevated temperature in yeast. Black-Right-Pointing-Pointer Tah18 was first identified as the yeast protein involved in NO synthesis. Black-Right-Pointing-Pointer Tah18-dependent NO synthesis confers tolerance to high-temperature on yeast cells. -- Abstract: Nitric oxide (NO) is a ubiquitous signaling molecule involved in the regulation of a large number of cellular functions. In the unicellular eukaryote yeast, NO may be involved in stress response pathways, but its role is poorly understood due to the lack of mammalian NO synthase (NOS) orthologues. Previously, we have proposed the oxidative stress-induced L-arginine synthesis and its physiologicalmore » role under stress conditions in yeast Saccharomyces cerevisiae. Here, our experimental results indicated that increased conversion of L-proline into L-arginine led to NO production in response to elevated temperature. We also showed that the flavoprotein Tah18, which was previously reported to transfer electrons to the Fe-S cluster protein Dre2, was involved in NO synthesis in yeast. Gene knockdown analysis demonstrated that Tah18-dependent NO synthesis confers high-temperature stress tolerance on yeast cells. As it appears that such a unique cell protection mechanism is specific to yeasts and fungi, it represents a promising target for antifungal activity.« less

The yeast genome may harbor hypoxia response elements (HRE).

PubMed

Ferreira, Túlio César; Hertzberg, Libi; Gassmann, Max; Campos, Elida Geralda

2007-01-01

The hypoxia-inducible factor-1 (HIF-1) is a heterodimeric transcription factor activated when cells are submitted to hypoxia. The heterodimer is composed of two subunits, HIF-1alpha and the constitutively expressed HIF-1beta. During normoxia, HIF-1alpha is degraded by the 26S proteasome, but hypoxia causes HIF-1alpha to be stabilized, enter the nucleus and bind to HIF-1beta, thus forming the active complex. The complex then binds to the regulatory sequences of various genes involved in physiological and pathological processes. The specific regulatory sequence recognized by HIF-1 is the hypoxia response element (HRE) that has the consensus sequence 5'BRCGTGVBBB3'. Although the basic transcriptional regulation machinery is conserved between yeast and mammals, Saccharomyces cerevisiae does not express HIF-1 subunits. However, we hypothesized that baker's yeast has a protein analogous to HIF-1 which participates in the response to changes in oxygen levels by binding to HRE sequences. In this study we screened the yeast genome for HREs using probabilistic motif search tools. We described 24 yeast genes containing motifs with high probability of being HREs (p-value<0.1) and classified them according to biological function. Our results show that S. cerevisiae may harbor HREs and indicate that a transcription factor analogous to HIF-1 may exist in this organism.

Discussion of teleomorphic and anamorphic Ascomycetous yeasts and yeast-like taxa

USDA-ARS?s Scientific Manuscript database

The relationship of ascomycetous yeasts with other members of the ascomycete fungi (Ascomycota) has been controversial for over 100 years. Because yeasts are morphologically simple, it was proposed that they represent primitive forms of ascomycetes (e.g., Guilliermond 1912). Alternatively, the ide...

Probiotic yeasts: Anti-inflammatory potential of various non-pathogenic strains in experimental colitis in mice

PubMed Central

Foligné, Benoît; Dewulf, Joëlle; Vandekerckove, Pascal; Pignède, Georges; Pot, Bruno

2010-01-01

AIM: To evaluate the in vitro immunomodulation capacity of various non-pathogenic yeast strains and to investigate the ability of some of these food grade yeasts to prevent experimental colitis in mice. METHODS: In vitro immunomodulation was assessed by measuring cytokines [interleukin (IL)-12p70, IL-10, tumor necrosis factor and interferon γ] released by human peripheral blood mononuclear cells after 24 h stimulation with 6 live yeast strains (Saccharomyces ssp.) and with bacterial reference strains. A murine model of acute 2-4-6-trinitrobenzene sulfonic acid (TNBS)-colitis was next used to evaluate the distinct prophylactic protective capacities of three yeast strains compared with the performance of prednisolone treatment. RESULTS: The six yeast strains all showed similar non-discriminating anti-inflammatory potential when tested on immunocompetent cells in vitro. However, although they exhibited similar colonization patterns in vivo, some yeast strains showed significant anti-inflammatory activities in the TNBS-induced colitis model, whereas others had weaker or no preventive effect at all, as evidenced by colitis markers (body-weight loss, macroscopic and histological scores, myeloperoxidase activities and blood inflammatory markers). CONCLUSION: A careful selection of strains is required among the biodiversity of yeasts for specific clinical studies, including applications in inflammatory bowel disease and other therapeutic uses. PMID:20440854

Pseudoephedrine/ephedrine shows potent anti-inflammatory activity against TNF-α-mediated acute liver failure induced by lipopolysaccharide/D-galactosamine.

PubMed

Wu, Zhongping; Kong, Xiangliang; Zhang, Tong; Ye, Jin; Fang, Zhaoqin; Yang, Xuejun

2014-02-05

The anti-inflammatory effects of pseudoephedrine/ephedrine were investigated using the experimental model of lipopolysaccharide (LPS)-induced acute liver failure in D-galactosamine (D-GalN)-sensitised male rats in order to elucidate effects other than sympathomimetic effects. Rats were intraperitoneally injected with D-GalN (400 mg/kg) and LPS (40 μg/kg) to induce acute liver failure. The treatment groups were then intraperitoneally administered pseudoephedrine/ephedrine at 0 h and 4 h after induction and the activation induced by treatment with pseudoephedrine and/or LPS on the primary Kupffer cells (KCs) was monitored. Compared with controls induced by GalN/LPS alone, pseudoephedrine dramatically reduced the infiltration of inflammatory cells and bile ductular hyperplasia and hepatic necrosis observed in liver sections. It inhibited both hepatocellular apoptosis and the expression of monocyte chemotactic protein-1. It lowered the production of tumour necrosis factor-α (TNF-α) in the beginning of acute liver failure induced by D-GalN/LPS. Correspondingly, levels of alanine aminotransferase (ALT), total bilirubin (TBIL) and malondialdehyde were attenuated. Ephedrine demonstrated all these identical protective effects as well. In addition, pseudoephedrine significantly suppressed the production of p-IκB-α, reducing the degradation of sequestered nuclear factor kappa B (NF-κB) in the cytoplasm, and inhibited the translocation of NF-κB/p65 to the nucleus, the transcription of TNF-α mRNA and the production of TNF-α in primary KCs. These results suggest that pseudoephedrine and ephedrine have a potent anti-inflammatory activity against D-GalN/LPS-induced acute liver failure in rats, and this comprehensive anti-inflammatory effect may result from the inhibition of TNF-α production. Copyright © 2013 Elsevier B.V. All rights reserved.

Nucleoside-modified mRNA vaccines induce potent T follicular helper and germinal center B cell responses.

PubMed

Pardi, Norbert; Hogan, Michael J; Naradikian, Martin S; Parkhouse, Kaela; Cain, Derek W; Jones, Letitia; Moody, M Anthony; Verkerke, Hans P; Myles, Arpita; Willis, Elinor; LaBranche, Celia C; Montefiori, David C; Lobby, Jenna L; Saunders, Kevin O; Liao, Hua-Xin; Korber, Bette T; Sutherland, Laura L; Scearce, Richard M; Hraber, Peter T; Tombácz, István; Muramatsu, Hiromi; Ni, Houping; Balikov, Daniel A; Li, Charles; Mui, Barbara L; Tam, Ying K; Krammer, Florian; Karikó, Katalin; Polacino, Patricia; Eisenlohr, Laurence C; Madden, Thomas D; Hope, Michael J; Lewis, Mark G; Lee, Kelly K; Hu, Shiu-Lok; Hensley, Scott E; Cancro, Michael P; Haynes, Barton F; Weissman, Drew

2018-06-04

T follicular helper (Tfh) cells are required to develop germinal center (GC) responses and drive immunoglobulin class switch, affinity maturation, and long-term B cell memory. In this study, we characterize a recently developed vaccine platform, nucleoside-modified, purified mRNA encapsulated in lipid nanoparticles (mRNA-LNPs), that induces high levels of Tfh and GC B cells. Intradermal vaccination with nucleoside-modified mRNA-LNPs encoding various viral surface antigens elicited polyfunctional, antigen-specific, CD4 + T cell responses and potent neutralizing antibody responses in mice and nonhuman primates. Importantly, the strong antigen-specific Tfh cell response and high numbers of GC B cells and plasma cells were associated with long-lived and high-affinity neutralizing antibodies and durable protection. Comparative studies demonstrated that nucleoside-modified mRNA-LNP vaccines outperformed adjuvanted protein and inactivated virus vaccines and pathogen infection. The incorporation of noninflammatory, modified nucleosides in the mRNA is required for the production of large amounts of antigen and for robust immune responses. © 2018 Pardi et al.

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.

High-Content Screening in Zebrafish Embryos Identifies Butafenacil as a Potent Inducer of Anemia

PubMed Central

Leet, Jessica K.; Lindberg, Casey D.; Bassett, Luke A.; Isales, Gregory M.; Yozzo, Krystle L.; Raftery, Tara D.; Volz, David C.

2014-01-01

Using transgenic zebrafish (fli1:egfp) that stably express enhanced green fluorescent protein (eGFP) within vascular endothelial cells, we recently developed and optimized a 384-well high-content screening (HCS) assay that enables us to screen and identify chemicals affecting cardiovascular development and function at non-teratogenic concentrations. Within this assay, automated image acquisition procedures and custom image analysis protocols are used to quantify body length, heart rate, circulation, pericardial area, and intersegmental vessel area within individual live embryos exposed from 5 to 72 hours post-fertilization. After ranking developmental toxicity data generated from the U.S. Environmental Protection Agency's (EPA's) zebrafish teratogenesis assay, we screened 26 of the most acutely toxic chemicals within EPA's ToxCast Phase-I library in concentration-response format (0.05–50 µM) using this HCS assay. Based on this screen, we identified butafenacil as a potent inducer of anemia, as exposure from 0.39 to 3.125 µM butafenacil completely abolished arterial circulation in the absence of effects on all other endpoints evaluated. Butafenacil is an herbicide that inhibits protoporphyrinogen oxidase (PPO) – an enzyme necessary for heme production in vertebrates. Using o-dianisidine staining, we then revealed that severe butafenacil-induced anemia in zebrafish was due to a complete loss of hemoglobin following exposure during early development. Therefore, six additional PPO inhibitors within the ToxCast Phase-I library were screened to determine whether anemia represents a common adverse outcome for these herbicides. Embryonic exposure to only one of these PPO inhibitors – flumioxazin – resulted in a similar phenotype as butafenacil, albeit not as severe as butafenacil. Overall, this study highlights the potential utility of this assay for (1) screening chemicals for cardiovascular toxicity and (2) prioritizing chemicals for future hypothesis

Adaptive response to chronic mild ethanol stress involves ROS, sirtuins and changes in chromosome dosage in wine yeasts.

PubMed

Adamczyk, Jagoda; Deregowska, Anna; Skoneczny, Marek; Skoneczna, Adrianna; Kwiatkowska, Aleksandra; Potocki, Leszek; Rawska, Ewa; Pabian, Sylwia; Kaplan, Jakub; Lewinska, Anna; Wnuk, Maciej

2016-05-24

Industrial yeast strains of economic importance used in winemaking and beer production are genomically diverse and subjected to harsh environmental conditions during fermentation. In the present study, we investigated wine yeast adaptation to chronic mild alcohol stress when cells were cultured for 100 generations in the presence of non-cytotoxic ethanol concentration. Ethanol-induced reactive oxygen species (ROS) and superoxide signals promoted growth rate during passages that was accompanied by increased expression of sirtuin proteins, Sir1, Sir2 and Sir3, and DNA-binding transcription regulator Rap1. Genome-wide array-CGH analysis revealed that yeast genome was shaped during passages. The gains of chromosomes I, III and VI and significant changes in the gene copy number in nine functional gene categories involved in metabolic processes and stress responses were observed. Ethanol-mediated gains of YRF1 and CUP1 genes were the most accented. Ethanol also induced nucleolus fragmentation that confirms that nucleolus is a stress sensor in yeasts. Taken together, we postulate that wine yeasts of different origin may adapt to mild alcohol stress by shifts in intracellular redox state promoting growth capacity, upregulation of key regulators of longevity, namely sirtuins and changes in the dosage of genes involved in the telomere maintenance and ion detoxification.

Metabolic analyses elucidate non-trivial gene targets for amplifying dihydroartemisinic acid production in yeast

PubMed Central

Misra, Ashish; Conway, Matthew F.; Johnnie, Joseph; Qureshi, Tabish M.; Lige, Bao; Derrick, Anne M.; Agbo, Eddy C.; Sriram, Ganesh

2013-01-01

Synthetic biology enables metabolic engineering of industrial microbes to synthesize value-added molecules. In this, a major challenge is the efficient redirection of carbon to the desired metabolic pathways. Pinpointing strategies toward this goal requires an in-depth investigation of the metabolic landscape of the organism, particularly primary metabolism, to identify precursor and cofactor availability for the target compound. The potent antimalarial therapeutic artemisinin and its precursors are promising candidate molecules for production in microbial hosts. Recent advances have demonstrated the production of artemisinin precursors in engineered yeast strains as an alternative to extraction from plants. We report the application of in silico and in vivo metabolic pathway analyses to identify metabolic engineering targets to improve the yield of the direct artemisinin precursor dihydroartemisinic acid (DHA) in yeast. First, in silico extreme pathway (ExPa) analysis identified NADPH-malic enzyme and the oxidative pentose phosphate pathway (PPP) as mechanisms to meet NADPH demand for DHA synthesis. Next, we compared key DHA-synthesizing ExPas to the metabolic flux distributions obtained from in vivo 13C metabolic flux analysis of a DHA-synthesizing strain. This comparison revealed that knocking out ethanol synthesis and overexpressing glucose-6-phosphate dehydrogenase in the oxidative PPP (gene YNL241C) or the NADPH-malic enzyme ME2 (YKL029C) are vital steps toward overproducing DHA. Finally, we employed in silico flux balance analysis and minimization of metabolic adjustment on a yeast genome-scale model to identify gene knockouts for improving DHA yields. The best strategy involved knockout of an oxaloacetate transporter (YKL120W) and an aspartate aminotransferase (YKL106W), and was predicted to improve DHA yields by 70-fold. Collectively, our work elucidates multiple non-trivial metabolic engineering strategies for improving DHA yield in yeast. PMID:23898325

The yeast Saccharomyces cerevisiae: an overview of methods to study autophagy progression.

PubMed

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

2015-03-01

Macroautophagy (hereafter autophagy) is a highly evolutionarily conserved process essential for sustaining cellular integrity, homeostasis, and survival. Most eukaryotic cells constitutively undergo autophagy at a low basal level. However, various stimuli, including starvation, organelle deterioration, stress, and pathogen infection, potently upregulate autophagy. The hallmark morphological feature of autophagy is the formation of the double-membrane vesicle known as the autophagosome. In yeast, flux through the pathway culminates in autophagosome-vacuole fusion, and the subsequent degradation of the resulting autophagic bodies and cargo by vacuolar hydrolases, followed by efflux of the breakdown products. Importantly, aberrant autophagy is associated with diverse human pathologies. Thus, there is a need for ongoing work in this area to further understand the cellular factors regulating this process. The field of autophagy research has grown exponentially in recent years, and although numerous model organisms are being used to investigate autophagy, the baker's yeast Saccharomyces cerevisiae remains highly relevant, as there are significant and unique benefits to working with this organism. In this review, we will focus on the current methods available to evaluate and monitor autophagy in S. cerevisiae, which in several cases have also been subsequently exploited in higher eukaryotes. Copyright © 2014 Elsevier Inc. All rights reserved.

Heavy ion-induced DNA double-strand breaks in yeast.

PubMed

Kiefer, Jürgen; Egenolf, Ralf; Ikpeme, Samuel

2002-02-01

Induction of DSBs in the diploid yeast, Saccharomyces cerevisiae, was measured by pulsed-field gel electrophoresis (PFGE) after the cells had been exposed on membrane filters to a variety of energetic heavy ions with values of linear energy transfer (LET) ranging from about 2 to 11,500 keV/microm, (241)Am alpha particles, and 80 keV X rays. After irradiation, the cells were lysed, and the chromosomes were separated by PFGE. The gels were stained with ethidium bromide, placed on a UV transilluminator, and analyzed using a computer-coupled camera. The fluorescence intensities of the larger bands were found to decrease exponentially with dose or particle fluence. The slope of this line corresponds to the cross section for at least one double-strand break (DSB), but closely spaced multiple breaks cannot be discriminated. Based on the known size of the native DNA molecules, breakage cross sections per base pair were calculated. They increased with LET until they reached a transient plateau value of about 6 x 10(-7) microm(2) at about 300-2000 keV/microm; they then rose for the higher LETs, probably reflecting the influence of delta electrons. The relative biological effectiveness for DNA breakage displays a maximum of about 2.5 around 100-200 keV/microm and falls below unity for LET values above 10(3) keV/microm. For these yeast cells, comparison of the derived breakage cross sections with the corresponding cross section for inactivation derived from the terminal slope of the survival curves shows a strong linear relationship between these cross sections, extending over several orders of magnitude.

Effect of Selenium on Lipid and Amino Acid Metabolism in Yeast Cells.

PubMed

Kieliszek, Marek; Błażejak, Stanisław; Bzducha-Wróbel, Anna; Kot, Anna M

2018-04-19

This article discusses the effect of selenium in aqueous solutions on aspects of lipid and amino acid metabolism in the cell biomass of Saccharomyces cerevisiae MYA-2200 and Candida utilis ATCC 9950 yeasts. The yeast biomass was obtained by using waste products (potato wastewater and glycerol). Selenium, at a dose of 20 mg/L of aqueous solution, affected the differentiation of cellular morphology. Yeast enriched with selenium was characterized by a large functional diversity in terms of protein and amino acid content. The protein content in the biomass of S. cerevisiae enriched with selenium (42.6%) decreased slightly as compared to that in the control sample without additional selenium supplementation (48.4%). Moreover, yeasts of both strains enriched with selenium contained a large amount of glutamic acid, aspartic acid, lysine, and leucine. Analysis of fatty acid profiles in S. cerevisiae yeast supplemented with selenium showed an increase in the unsaturated fatty acid content (e.g., C18:1). The presence of margaric acid (C17:0) and hexadecanoic acid (C17:1) was found in the C. utilis biomass enriched with selenium, in contrast to that of S. cerevisiae. These results indicate that selenium may induce lipid peroxidation, which consequently affects the loss of integrity of the cytoplasmic membrane. Yeast enriched with selenium with optimal amino acid and lipid composition can be used to prepare a novel formula of dietary supplements, which can be applied directly to various diets for both humans and animals.

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.

Cloning of a yeast alpha-amylase promoter and its regulated heterologous expression

DOEpatents

Gao, Johnway [Richland, WA; Skeen, Rodney S [Pendleton, OR; Hooker, Brian S [Kennewick, WA; Anderson, Daniel B [Pasco, WA

2003-04-01

The present invention provides the promoter clone discovery of an alpha-amylase gene of a starch utilizing yeast strain Schwanniomyces castellii. The isolated alpha-amylase promoter is an inducible promoter, which can regulate strong gene expression in starch culture medium.

Boletus edulis ribonucleic acid - a potent apoptosis inducer in human colon adenocarcinoma cells.

PubMed

Lemieszek, Marta Kinga; Ribeiro, Miguel; Guichard Alves, Helena; Marques, Guilhermina; Nunes, Fernando Milheiro; Rzeski, Wojciech

2016-07-13

Despite the large popularity of the Boletus edulis mushroom, little is known about its influence on human health and the possibilities of its therapeutic use. Nevertheless, several reports revealed the usefulness of biopolymers isolated from it in cancer treatment. Our previous studies have shown that B. edulis water soluble biopolymers are not toxic against normal colon epithelial cells (CCD841 CoTr) and at the same concentration range elicited a very prominent antiproliferative effect in colon cancer cells (LS180) which was accompanied with cell cycle arrest in the G0/G1 phase. The purpose of the present study was to verify the proapoptotic properties of a selected fraction from B. edulis - BE3, as well as determine its chemical nature. The BE3 fraction was extracted with hot water and purified by anion-exchange chromatography. Further chemical examinations revealed that BE3 consists mainly of ribonucleic acid (59.1%). The ability of BE3 to induce programmed cell death was examined in human colon cancer cell lines LS180 and HT-29 by measuring caspase activation, DNA fragmentation and expression of BAX, BCL2, TP53 and CDKN1A genes. The sensitivity of colon cancer cells with silenced BAX, TP53 and CDKN1A expression to BE3 treatment was also evaluated. We have demonstrated for the first time that the BE3 fraction is a potent apoptosis inducer in human colon cancer cells. The revealed mechanism of apoptosis triggering was dependent on the presence of functional p53 and consequently was a little different in investigated cell lines. Our results indicated that BE3 stimulated proapoptotic genes BAX (LS180, HT-29), TP53 (LS180) and CDKN1A (HT-29) while at the same time silenced the expression of the key prosurvival gene BCL2 (LS180, HT-29). The obtained results indicate the high therapeutic potential of the BE3 fraction against colon cancer, yet it is necessary to further confirm fraction efficacy and safety in animal and clinical studies.

[Thermoresistance in Saccharomyces cerevisiae yeasts].

PubMed

Kaliuzhin, V A

2011-01-01

Under natural conditions, yeast Saccharomyces cerevisiae reproduce, as a rule, on the surface of solid or liquid medium. Thus, life cycle of yeast populations is substantially influenced by diurnal changes in ambient temperature. The pattern in the response of unrestricted yeast S. cerevisiae culture to changes in the temperature of cultivation is revealed experimentally. Yeast population, in the absence of environmental constraints on the functioning of cell chemosmotic bioenergetic system, demonstrates the ability of thermoresistance when the temperature of cultivation switches from the range of 12-36 degrees C to 37.5-40 degrees C. During the transient period that is associated with the temperature switching and lasts from 1 to 4 turnover cycles, yeast reproduction rate remains 1.5-2 times higher than under stationary conditions. This is due to evolutionary acquired adaptive activity of cell chemosmotic system. After the adaptive resources exhausting, yeast thermoresistance fully recovers at the temperature range of 12-36 degrees C within one generation time under conditions of both restricted and unrestricted nourishment. Adaptive significance of such thermoresistance seems obvious enough--it allows maintaining high reproduction rate in yeast when ambient temperature is reaching a brief maximum shortly after noon.

Interaction Between Yeasts and Zinc

NASA Astrophysics Data System (ADS)

Nicola, Raffaele De; Walker, Graeme

Zinc is an essential trace element in biological systems. For example, it acts as a cellular membrane stabiliser, plays a critical role in gene expression and genome modification and activates nearly 300 enzymes, including alcohol dehydrogenase. The present chapter will be focused on the influence of zinc on cell physiology of industrial yeast strains of Saccharomyces cerevisiae, with special regard to the uptake and subsequent utilisation of this metal. Zinc uptake by yeast is metabolism-dependent, with most of the available zinc translocated very quickly into the vacuole. At cell division, zinc is distributed from mother to daughter cells and this effectively lowers the individual cellular zinc concentration, which may become zinc depleted at the onset of the fermentation. Zinc influences yeast fermentative performance and examples will be provided relating to brewing and wine fermentations. Industrial yeasts are subjected to several stresses that may impair fermentation performance. Such stresses may also impact on yeast cell zinc homeostasis. This chapter will discuss the practical implications for the correct management of zinc bioavailability for yeast-based biotechnologies aimed at improving yeast growth, viability, fermentation performance and resistance to environmental stresses

21 CFR 172.896 - Dried yeasts.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Dried yeasts. 172.896 Section 172.896 Food and... PERMITTED FOR DIRECT ADDITION TO FOOD FOR HUMAN CONSUMPTION Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis...

Heterologous expression of cytotoxic sesquiterpenoids from the medicinal mushroom Lignosus rhinocerotis in yeast.

PubMed

Yap, Hui-Yeng Yeannie; Muria-Gonzalez, Mariano Jordi; Kong, Boon-Hong; Stubbs, Keith A; Tan, Chon-Seng; Ng, Szu-Ting; Tan, Nget-Hong; Solomon, Peter S; Fung, Shin-Yee; Chooi, Yit-Heng

2017-06-12

Genome mining facilitated by heterologous systems is an emerging approach to access the chemical diversity encoded in basidiomycete genomes. In this study, three sesquiterpene synthase genes, GME3634, GME3638, and GME9210, which were highly expressed in the sclerotium of the medicinal mushroom Lignosus rhinocerotis, were cloned and heterologously expressed in a yeast system. Metabolite profile analysis of the yeast culture extracts by GC-MS showed the production of several sesquiterpene alcohols (C 15 H 26 O), including cadinols and germacrene D-4-ol as major products. Other detected sesquiterpenes include selina-6-en-4-ol, β-elemene, β-cubebene, and cedrene. Two purified major compounds namely (+)-torreyol and α-cadinol synthesised by GME3638 and GME3634 respectively, are stereoisomers and their chemical structures were confirmed by 1 H and 13 C NMR. Phylogenetic analysis revealed that GME3638 and GME3634 are a pair of orthologues, and are grouped together with terpene synthases that synthesise cadinenes and related sesquiterpenes. (+)-Torreyol and α-cadinol were tested against a panel of human cancer cell lines and the latter was found to exhibit selective potent cytotoxicity in breast adenocarcinoma cells (MCF7) with IC 50 value of 3.5 ± 0.58 μg/ml while α-cadinol is less active (IC 50  = 18.0 ± 3.27 μg/ml). This demonstrates that yeast-based genome mining, guided by transcriptomics, is a promising approach for uncovering bioactive compounds from medicinal mushrooms.

PCR-based method for the rapid identification of astaxanthin-accumulating yeasts (Phaffia spp.).

PubMed

Colabella, Fernando; Libkind, Diego

2016-01-01

It has been recently found that the natural distribution, habitat, and genetic diversity of astaxanthin-producing yeasts (i.e. Phaffia rhodozyma, synonym Xanthophyllomyces dendrorhous) is much greater than previously thought. P. rhodozyma is biotechnologically exploited due to its ability to produce the carotenoid pigment astaxanthin and thus, it is used as a natural source of this pigment for aquaculture. P. rhodozyma was also capable of synthesizing the potent UVB sunscreen mycosporine-glutaminol-glucoside (MGG). Therefore, further environmental studies are needed to elucidate its ecological aspects and detect new potential strains for the production of astaxanthin and MGG. However, obtaining new isolates of P. rhodozyma and related species is not always easy due to its low abundance and the presence of other sympatric and pigmented yeasts. In this work we report a successful development of a species-specific primer which has the ability to quickly and accurately detecting isolates representing all known lineages of the genus Phaffia (including novel species of the genus) and excluding closely related taxa. For this purpose, a primer of 20 nucleotides (called PhR) was designed to be used in combination with universal primers ITS3 and NL4 in a multiplex amplification. The proposed method has the sensitivity and specificity required for the precise detection of new isolates, and therefore represents an important tool for the environmental search for novel astaxanthin-producing yeasts. Copyright © 2015 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.

Homocysteine regulates fatty acid and lipid metabolism in yeast.

PubMed

Visram, Myriam; Radulovic, Maja; Steiner, Sabine; Malanovic, Nermina; Eichmann, Thomas O; Wolinski, Heimo; Rechberger, Gerald N; Tehlivets, Oksana

2018-04-13

S -Adenosyl-l-homocysteine hydrolase (AdoHcy hydrolase; Sah1 in yeast/AHCY in mammals) degrades AdoHcy, a by-product and strong product inhibitor of S -adenosyl-l-methionine (AdoMet)-dependent methylation reactions, to adenosine and homocysteine (Hcy). This reaction is reversible, so any elevation of Hcy levels, such as in hyperhomocysteinemia (HHcy), drives the formation of AdoHcy, with detrimental consequences for cellular methylation reactions. HHcy, a pathological condition linked to cardiovascular and neurological disorders, as well as fatty liver among others, is associated with a deregulation of lipid metabolism. Here, we developed a yeast model of HHcy to identify mechanisms that dysregulate lipid metabolism. Hcy supplementation to wildtype cells up-regulated cellular fatty acid and triacylglycerol content and induced a shift in fatty acid composition, similar to changes observed in mutants lacking Sah1. Expression of the irreversible bacterial pathway for AdoHcy degradation in yeast allowed us to dissect the impact of AdoHcy accumulation on lipid metabolism from the impact of elevated Hcy. Expression of this pathway fully suppressed the growth deficit of sah1 mutants as well as the deregulation of lipid metabolism in both the sah1 mutant and Hcy-exposed wildtype, showing that AdoHcy accumulation mediates the deregulation of lipid metabolism in response to elevated Hcy in yeast. Furthermore, Hcy supplementation in yeast led to increased resistance to cerulenin, an inhibitor of fatty acid synthase, as well as to a concomitant decline of condensing enzymes involved in very long-chain fatty acid synthesis, in line with the observed shift in fatty acid content and composition. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Homocysteine regulates fatty acid and lipid metabolism in yeast

PubMed Central

Visram, Myriam; Radulovic, Maja; Steiner, Sabine; Malanovic, Nermina; Eichmann, Thomas O.; Wolinski, Heimo; Rechberger, Gerald N.; Tehlivets, Oksana

2018-01-01

S-Adenosyl-l-homocysteine hydrolase (AdoHcy hydrolase; Sah1 in yeast/AHCY in mammals) degrades AdoHcy, a by-product and strong product inhibitor of S-adenosyl-l-methionine (AdoMet)-dependent methylation reactions, to adenosine and homocysteine (Hcy). This reaction is reversible, so any elevation of Hcy levels, such as in hyperhomocysteinemia (HHcy), drives the formation of AdoHcy, with detrimental consequences for cellular methylation reactions. HHcy, a pathological condition linked to cardiovascular and neurological disorders, as well as fatty liver among others, is associated with a deregulation of lipid metabolism. Here, we developed a yeast model of HHcy to identify mechanisms that dysregulate lipid metabolism. Hcy supplementation to wildtype cells up-regulated cellular fatty acid and triacylglycerol content and induced a shift in fatty acid composition, similar to changes observed in mutants lacking Sah1. Expression of the irreversible bacterial pathway for AdoHcy degradation in yeast allowed us to dissect the impact of AdoHcy accumulation on lipid metabolism from the impact of elevated Hcy. Expression of this pathway fully suppressed the growth deficit of sah1 mutants as well as the deregulation of lipid metabolism in both the sah1 mutant and Hcy-exposed wildtype, showing that AdoHcy accumulation mediates the deregulation of lipid metabolism in response to elevated Hcy in yeast. Furthermore, Hcy supplementation in yeast led to increased resistance to cerulenin, an inhibitor of fatty acid synthase, as well as to a concomitant decline of condensing enzymes involved in very long-chain fatty acid synthesis, in line with the observed shift in fatty acid content and composition. PMID:29414770

Components of a Fanconi-like pathway control Pso2-independent DNA interstrand crosslink repair in yeast.

PubMed

Ward, Thomas A; Dudášová, Zuzana; Sarkar, Sovan; Bhide, Mangesh R; Vlasáková, Danuša; Chovanec, Miroslav; McHugh, Peter J

2012-01-01

Fanconi anemia (FA) is a devastating genetic disease, associated with genomic instability and defects in DNA interstrand cross-link (ICL) repair. The FA repair pathway is not thought to be conserved in budding yeast, and although the yeast Mph1 helicase is a putative homolog of human FANCM, yeast cells disrupted for MPH1 are not sensitive to ICLs. Here, we reveal a key role for Mph1 in ICL repair when the Pso2 exonuclease is inactivated. We find that the yeast FANCM ortholog Mph1 physically and functionally interacts with Mgm101, a protein previously implicated in mitochondrial DNA repair, and the MutSα mismatch repair factor (Msh2-Msh6). Co-disruption of MPH1, MGM101, MSH6, or MSH2 with PSO2 produces a lesion-specific increase in ICL sensitivity, the elevation of ICL-induced chromosomal rearrangements, and persistence of ICL-associated DNA double-strand breaks. We find that Mph1-Mgm101-MutSα directs the ICL-induced recruitment of Exo1 to chromatin, and we propose that Exo1 is an alternative 5'-3' exonuclease utilised for ICL repair in the absence of Pso2. Moreover, ICL-induced Rad51 chromatin loading is delayed when both Pso2 and components of the Mph1-Mgm101-MutSα and Exo1 pathway are inactivated, demonstrating that the homologous recombination stages of ICL repair are inhibited. Finally, the FANCJ- and FANCP-related factors Chl1 and Slx4, respectively, are also components of the genetic pathway controlled by Mph1-Mgm101-MutSα. Together this suggests that a prototypical FA-related ICL repair pathway operates in budding yeast, which acts redundantly with the pathway controlled by Pso2, and is required for the targeting of Exo1 to chromatin to execute ICL repair.

Barcode Sequencing Screen Identifies SUB1 as a Regulator of Yeast Pheromone Inducible Genes

PubMed Central

Sliva, Anna; Kuang, Zheng; Meluh, Pamela B.; Boeke, Jef D.

2016-01-01

The yeast pheromone response pathway serves as a valuable model of eukaryotic mitogen-activated protein kinase (MAPK) pathways, and transcription of their downstream targets. Here, we describe application of a screening method combining two technologies: fluorescence-activated cell sorting (FACS), and barcode analysis by sequencing (Bar-Seq). Using this screening method, and pFUS1-GFP as a reporter for MAPK pathway activation, we readily identified mutants in known mating pathway components. In this study, we also include a comprehensive analysis of the FUS1 induction properties of known mating pathway mutants by flow cytometry, featuring single cell analysis of each mutant population. We also characterized a new source of false positives resulting from the design of this screen. Additionally, we identified a deletion mutant, sub1Δ, with increased basal expression of pFUS1-GFP. Here, in the first ChIP-Seq of Sub1, our data shows that Sub1 binds to the promoters of about half the genes in the genome (tripling the 991 loci previously reported), including the promoters of several pheromone-inducible genes, some of which show an increase upon pheromone induction. Here, we also present the first RNA-Seq of a sub1Δ mutant; the majority of genes have no change in RNA, but, of the small subset that do, most show decreased expression, consistent with biochemical studies implicating Sub1 as a positive transcriptional regulator. The RNA-Seq data also show that certain pheromone-inducible genes are induced less in the sub1Δ mutant relative to the wild type, supporting a role for Sub1 in regulation of mating pathway genes. The sub1Δ mutant has increased basal levels of a small subset of other genes besides FUS1, including IMD2 and FIG1, a gene encoding an integral membrane protein necessary for efficient mating. PMID:26837954

21 CFR 172.896 - Dried yeasts.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Dried yeasts. 172.896 Section 172.896 Food and... Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis) may be safely used in food provided the total folic acid...

21 CFR 172.896 - Dried yeasts.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Dried yeasts. 172.896 Section 172.896 Food and... Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis) may be safely used in food provided the total folic acid...

21 CFR 172.896 - Dried yeasts.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Dried yeasts. 172.896 Section 172.896 Food and... Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis) may be safely used in food provided the total folic acid...

21 CFR 172.896 - Dried yeasts.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Dried yeasts. 172.896 Section 172.896 Food and... Multipurpose Additives § 172.896 Dried yeasts. Dried yeast (Saccharomyces cerevisiae and Saccharomyces fragilis) and dried torula yeast (Candida utilis) may be safely used in food provided the total folic acid...

The effect of newly induced mutations on the fitness of genotypes and populations of yeast (Saccharomyces cerevisiae).

PubMed

Orthen, E; Lange, P; Wöhrmann, K

1984-12-01

This paper analyses the fate of artificially induced mutations and their importance to the fitness of populations of the yeast, Saccharomyces cerevisiae, an increasingly important model organism in population genetics. Diploid strains, treated with UV and EMS, were cultured asexually for approximately 540 generations and under conditions where the asexual growth was interrupted by a sexual phase. Growth rates of 100 randomly sampled diploid clones were estimated at the beginning and at the end of the experiment. After the induction of sporulation the growth rates of 100 randomly sampled spores were measured. UV and EMS treatment decreases the average growth rate of the clones significantly but increases the variability in comparison to the untreated control. After selection over approximately 540 generations, variability in growth rates was reduced to that of the untreated control. No increase in mean population fitness was observed. However, the results show that after selection there still exists a large amount of hidden genetic variability in the populations which is revealed when the clones are cultivated in environments other than those in which selection took place. A sexual phase increased the reduction of the induced variability.

Screening and characterization of amylase and cellulase activities in psychrotolerant yeasts.

PubMed

Carrasco, Mario; Villarreal, Pablo; Barahona, Salvador; Alcaíno, Jennifer; Cifuentes, Víctor; Baeza, Marcelo

2016-02-19

Amylases and cellulases have great potential for application in industries such as food, detergent, laundry, textile, baking and biofuels. A common requirement in these fields is to reduce the temperatures of the processes, leading to a continuous search for microorganisms that secrete cold-active amylases and cellulases. Psychrotolerant yeasts are good candidates because they inhabit cold-environments. In this work, we analyzed the ability of yeasts isolated from the Antarctic region to grow on starch or carboxymethylcellulose, and their potential extracellular amylases and cellulases. All tested yeasts were able to grow with soluble starch or carboxymethylcellulose as the sole carbon source; however, not all of them produced ethanol by fermentation of these carbon sources. For the majority of the yeast species, the extracellular amylase or cellulase activity was higher when cultured in medium supplemented with glucose rather than with soluble starch or carboxymethylcellulose. Additionally, higher amylase activities were observed when tested at pH 5.4 and 6.2, and at 30-37 °C, except for Rhodotorula glacialis that showed elevated activity at 10-22 °C. In general, cellulase activity was high until pH 6.2 and between 22-37 °C, while the sample from Mrakia blollopis showed high activity at 4-22 °C. Peptide mass fingerprinting analysis of a potential amylase from Tetracladium sp. of about 70 kDa, showed several peptides with positive matches with glucoamylases from other fungi. Almost all yeast species showed extracellular amylase or cellulase activity, and an inducing effect by the respective substrate was observed in a minor number of yeasts. These enzymatic activities were higher at 30 °C in most yeast, with highest amylase and cellulase activity in Tetracladium sp. and M. gelida, respectively. However, Rh. glacialis and M. blollopis displayed high amylase or cellulase activity, respectively, under 22 °C. In this sense, these yeasts are interesting

Novel endophytic yeast Rhodotorula mucilaginosa strain PTD3 II: production of xylitol and ethanol in the presence of inhibitors.

PubMed

Vajzovic, Azra; Bura, Renata; Kohlmeier, Kevin; Doty, Sharon L

2012-10-01

A systematic study was conducted characterizing the effect of furfural, 5-hydroxymethylfurfural (5-HMF), and acetic acid concentration on the production of xylitol and ethanol by a novel endophytic yeast, Rhodotorula mucilaginosa strain PTD3. The influence of different inhibitor concentrations on the growth and fermentation abilities of PTD3 cultivated in synthetic nutrient media containing 30 g/l xylose or glucose were measured during liquid batch cultures. Concentrations of up to 5 g/l of furfural stimulated production of xylitol to 77 % of theoretical yield (10 % higher compared to the control) by PTD3. Xylitol yields produced by this yeast were not affected in the presence of 5-HMF at concentrations of up to 3 g/l. At higher concentrations of furfural and 5-HMF, xylitol and ethanol yields were negatively affected. The higher the concentration of acetic acid present in a media, the higher the ethanol yield approaching 99 % of theoretical yield (15 % higher compared to the control) was produced by the yeast. At all concentrations of acetic acid tested, xylitol yield was lowered. PTD3 was capable of metabolizing concentrations of 5, 15, and 5 g/l of furfural, 5-HMF, and acetic acid, respectively. This yeast would be a potent candidate for the bioconversion of lignocellulosic sugars to biochemicals given that in the presence of low concentrations of inhibitors, its xylitol and ethanol yields are stimulated, and it is capable of metabolizing pretreatment degradation products.

Targeting the Oxidative Stress Response System of Fungi with Redox-Potent Chemosensitizing Agents

PubMed Central

Kim, Jong H.; Chan, Kathleen L.; Faria, Natália C. G.; Martins, M. de L.; Campbell, Bruce C.

2012-01-01

The cellular antioxidant system is a target in the antifungal action of amphotericin B (AMB) and itraconazole (ITZ), in filamentous fungi. The sakAΔ mutant of Aspergillus fumigatus, a mitogen-activated protein kinase (MAPK) gene deletion mutant in the antioxidant system, was found to be more sensitive to AMB or ITZ than other A. fumigatus strains, a wild type and a mpkCΔ mutant (a MAPK gene deletion mutant in the polyalcohol sugar utilization system). Complete fungal kill (≥99.9%) by ITZ or AMB was also achieved by much lower dosages for the sakAΔ mutant than for the other strains. It appears msnA, an Aspergillus ortholog to Saccharomyces cerevisiae MSN2 (encoding a stress-responsive C2H2-type zinc-finger regulator) and sakA and/or mpkC (upstream MAPKs) are in the same stress response network under tert-butyl hydroperoxide (t-BuOOH)-, hydrogen peroxide (H2O2)- or AMB-triggered toxicity. Of note is that ITZ-sensitive yeast pathogens were also sensitive to t-BuOOH, showing a connection between ITZ sensitivity and antioxidant capacity of fungi. Enhanced antifungal activity of AMB or ITZ was achieved when these drugs were co-applied with redox-potent natural compounds, 2,3-dihydroxybenzaldehyde, thymol or salicylaldehyde, as chemosensitizing agents. We concluded that redox-potent compounds, which target the antioxidant system in fungi, possess a chemosensitizing capacity to enhance efficacy of conventional drugs. PMID:22438852

[Clone, construct, expression and verification of lactoferricin B gene and several sequence mutations in yeast].

PubMed

Feng, Yong-qian; Zha, Xiao-jun; Zhai, Chao-yang

2007-07-01

To construct the eucaryotic recombinant plasmid of pYES2/LactoferricinB expressing in yeast of S. cerevisiae, of which the expressed protein antibacterial activity was verified in preliminary. By self-template PCR method, the gene of Lactoferricin B and its several sequence mutations were amplified with the parts of the pre-synthesized single chains. And then Lactoferricin B gene and its mutants were cloned into the vector of pYES2 to construct the recombined expression plasmid pYES2/Lactoferricin B etc. extracted and used to transform the yeast S. cerevisiae. The expressions of proteins were determined after induced by galactose. The expression proteins were collected and purified by hydronium-exchange column, and the bacterial inhibited test was applied to identify the protein antibacterial activities. The PCR amplifying and DNA sequencing tests indicated that the purpose plasmid contained the Lactoferricin B gene and several mutations. The induced target proteins were confirmed by SDS-PAGE electrophoresis and mass spectrum test. The protein antibacterial activities of mutations were verified in preliminary. The recombined plasmid pYES2/Lactoferricin B etc. are successfully constructed and induced to express in yeast cell of S. cerevisiae; the obtained recombined protein of Lactoferricin B provides a basis for further research work on the biological function and antibacterial activity.

Biostimulation effects of low-energy laser radiation on yeast cell suspensions

NASA Astrophysics Data System (ADS)

Anghel, Sorin; Stanescu, Constantin S.; Giosanu, Dana; Neagu, Ionica; Savulescu, Geta; Iorga-Siman, Ion

2000-02-01

This paper presents work to determine the effects produced by low energy laser radiation on the metabolism and growth of a yeast cell suspension. As experimental material, we used young yeast culture in liquid medium, then distributed on a solid medium, to obtain isolated colonies. As laser source, we used a He-Ne laser, and the irradiation was made with different exposure times. Form each irradiated material, a sample of white grape sterile must was sowed, that has fermented at 18 divided by 20 degrees C for 10 divided by 15 days, after that some properties was tested. Some microscopic studies were also made. The results prove some influence of low energy laser irradiation, which can induce mutations, with new properties of the irradiated material. These mutations can be obtained in a positive sense, with new and important perspectives in wine industry. Also, we observed an inhibitory effect of the laser radiation on the yeast cell growth, due, probably to the too high values of the exposure.

Prevalence of HBsAg and high-risk behaviors in pregnant women rerring to Urban Health Centers in Isfahan province

PubMed Central

Ahmadi, Mahboubeh; Toghyani, Raheleh; Shahidi, Shahla; Izadi, Minoo; Merasi, Mohammad Reza; Agdak, Pejman; Meshkaty, Marjan; Nikkhahfard, Mojghan

2011-01-01

BACKGROUND: About 5% of the world population are carriers of the hepatitis B virus which is not the same in different areas of the world. Iran, with a rate of 2-3%, is among the countries with average prevalence. The aim of this study was to determine the prevalence of HBsAg and its associated factors in pregnant women who referred to urban health centers in Isfahan Province. METHODS: This was a descriptive study conducted on 1078 pregnant women who had referred to the urban health centers of Isfahan, Borkhar, Meymeh, Khomeini Shahr, Lenjan and Najaf Abad in 2009 in order to register their physical condition and receive pregnancy care. Random sampling method by quota was done. First, a questionnaire including demographic characteristics and history of high-risk behaviors in mothers and their husbands was completed. Then, a blood sample was taken and evaluated for the HB virus surface antigen. Finally, the data was analyzed using SPSS software, Chi-square, Fisher and Logistic Regression tests. RESULTS: HB virus surface antigen was traced in the serum of 0.5% of the participants. The average age of subjects was 26.1± 4.9 (mean±sd) years old and the average pregnancy number was 1.79 + 1.0 (mean±sd). Using Fisher's test as well as logistic regression test and based on the previous history of high-risk sexual behaviors and tattooing, a significant difference was observed between the two groups with positive and negative HBsAg (p ≤ 0.05). CONCLUSIONS: Instructing the target groups and close monitoring of the high-risk centers such as beauty shops, tattooing centers and etc, and also staff training on how to use disposable and sterilized equipments have to be accomplished at the right time. PMID:22039379

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.

Immunogenicity of transgenic plant-derived hepatitis B surface antigen.

PubMed Central

Thanavala, Y; Yang, Y F; Lyons, P; Mason, H S; Arntzen, C

1995-01-01

The focus of the Children's Vaccine Initiative is to encourage the discovery of technology that will make vaccines more readily available to developing countries. Our strategy has been to genetically engineer plants so that they can be used as inexpensive alternatives to fermentation systems for production of subunit antigens. In this paper we report on the immunological response elicited in vivo by using recombinant hepatitis B surface antigen (rHBsAg) purified from transgenic tobacco leaves. The anti-hepatitis B response to the tobacco-derived rHBsAg was qualitatively similar to that obtained by immunizing mice with yeast-derived rHBsAg (commercial vaccine). Additionally, T cells obtained from mice primed with the tobacco-derived rHBsAg could be stimulated in vitro by the tobacco-derived rHBsAg, yeast-derived rHBsAg, and by a synthetic peptide that represents part of the a determinant located in the S region (139-147) of HBsAg. Further support for the integrity of the T-cell epitope of the tobacco-derived rHBsAg was obtained by testing the ability of the primed T cells to proliferate in vitro after stimulation with a monoclonal anti-idiotype and an anti-idiotype-derived peptide, both of which mimic the group-specific a determinant of HBsAg. In total, we have conclusively demonstrated that both B- and T-cell epitopes of HBsAg are preserved when the antigen is expressed in a transgenic plant. PMID:7724566

Preparation and testing of a Haemophilus influenzae Type b/Hepatitis B surface antigen conjugate vaccine.

PubMed

An, So Jung; Woo, Joo Sung; Chae, Myung Hwa; Kothari, Sudeep; Carbis, Rodney

2015-03-24

The majority of conjugate vaccines focus on inducing an antibody response to the polysaccharide antigen and the carrier protein is present primarily to induce a T-cell dependent response. In this study conjugates consisting of poly(ribosylribitolphosphate) (PRP) purified from Haemophilus influenzae Type b bound to Hepatitis B virus surface antigen (HBsAg) virus like particles were prepared with the aim of inducing an antibody response to not only the PRP but also the HBsAg. A conjugate consisting of PRP bound to HBsAg via an adipic acid dihydrazide (ADH) spacer induced strong IgG antibodies to both the PRP and HBsAg. When conjugation was performed without the ADH spacer the induction of an anti-PRP response was equivalent to that seen by conjugate with the ADH spacer, however, a negligible anti-HBsAg response was induced. For comparison, PRP was conjugated to diphtheria toxoid (DT) and Vi polysaccharide purified from Salmonella Typhi conjugated to HBsAg both using an ADH spacer. The PRPAH-DT conjugate induced strong anti-PRP and anti-DT responses, the Vi-AHHBsAg conjugate induced a good anti-HBsAg response but not as strong as that induced by the PRPAH-HBsAg conjugate. This study demonstrated that in mice it was possible to induce robust antibody responses to both polysaccharide and carrier protein provided the conjugate has certain physico-chemical properties. A PRPAH-HBsAg conjugate with the capacity to induce anti-PRP and anti-HBsAg responses could be incorporated into a multivalent pediatric vaccine and simplify formulation of such a vaccine. Copyright © 2015 Elsevier Ltd. All rights reserved.

Characterization of the "viable but nonculturable" (VBNC) state in the wine spoilage yeast Brettanomyces.

PubMed

Serpaggi, Virginie; Remize, Fabienne; Recorbet, Ghislaine; Gaudot-Dumas, Eliane; Sequeira-Le Grand, Anabelle; Alexandre, Hervé

2012-06-01

Although the viable but not culturable (VBNC) state has been studied in detail in bacteria, it has been suggested that maintenance of viability with loss of culturability also exists in eukaryotic cells, such as in the wine spoilage yeast Brettanomyces. To provide conclusive evidence for the existence of a VBNC state in this yeast, we investigated its capacity to become viable and nonculturable after sulfite stress, and its ability to recover culturability after stressor removal. Sulfite addition induced loss of culturability but maintenance of viability. Increasing the medium pH to decrease the concentration of toxic SO(2) allowed yeast cells to become culturable again, thus demonstrating the occurrence of a VBNC state in Brettanomyces upon SO(2) exposure. Relative to culturable Brettanomyces, VBNC yeast cells were found to display a 22% decrease in size on the basis of laser granulometry. Assays for 4-ethylguaiacol and 4-ethylphenol, volatile phenols produced by Brettanomyces, indicated that spoilage compound production could persist in VBNC cells. These morphological and physiological changes in VBNC Brettanomyces were coupled to extensive protein pattern modifications, as inferred by comparative two-dimensional electrophoresis and mass spectrometric analyses. Upon identification of 53 proteins out of the 168 spots whose abundance was significantly modified in treated cells relative to control, we propose that the SO(2)-induced VBNC state in Brettanomyces is characterized by a reduced glycolytic flux coupled to changes in redox homeostatis/protein turnover-related processes. This study points out the existence of common mechanisms between yeast and bacteria upon entry to the VBNC state. Copyright © 2012 Elsevier Ltd. All rights reserved.

Brewing characteristics of piezosensitive sake yeasts

NASA Astrophysics Data System (ADS)

Nomura, Kazuki; Hoshino, Hirofumi; Igoshi, Kazuaki; Onozuka, Haruka; Tanaka, Erika; Hayashi, Mayumi; Yamazaki, Harutake; Takaku, Hiroaki; Iguchi, Akinori; Shigematsu, Toru

2018-04-01

Application of high hydrostatic pressure (HHP) treatment to food processing is expected as a non-thermal fermentation regulation technology that supresses over fermentation. However, the yeast Saccharomyces cerevisiae used for Japanese rice wine (sake) brewing shows high tolerance to HHP. Therefore, we aimed to generate pressure-sensitive (piezosensitive) sake yeast strains by mating sake with piezosensitive yeast strains to establish an HHP fermentation regulation technology and extend the shelf life of fermented foods. The results of phenotypic analyses showed that the generated yeast strains were piezosensitive and exhibited similar fermentation ability compared with the original sake yeast strain. In addition, primary properties of sake brewed using these strains, such as ethanol concentration, sake meter value and sake flavor compounds, were almost equivalent to those obtained using the sake yeast strain. These results suggest that the piezosensitive strains exhibit brewing characteristics essentially equivalent to those of the sake yeast strain.

Discovery of plant extracts that greatly delay yeast chronological aging and have different effects on longevity-defining cellular processes

PubMed Central

Samson, Eugenie; Arlia-Ciommo, Anthony; Dakik, Pamela; Cortes, Berly; Feldman, Rachel; Mohtashami, Sadaf; McAuley, Mélissa; Chancharoen, Marisa; Rukundo, Belise; Simard, Éric; Titorenko, Vladimir I.

2016-01-01

We discovered six plant extracts that increase yeast chronological lifespan to a significantly greater extent than any of the presently known longevity-extending chemical compounds. One of these extracts is the most potent longevity-extending pharmacological intervention yet described. We show that each of the six plant extracts is a geroprotector which delays the onset and decreases the rate of yeast chronological aging by eliciting a hormetic stress response. We also show that each of these extracts has different effects on cellular processes that define longevity in organisms across phyla. These effects include the following: 1) increased mitochondrial respiration and membrane potential; 2) augmented or reduced concentrations of reactive oxygen species; 3) decreased oxidative damage to cellular proteins, membrane lipids, and mitochondrial and nuclear genomes; 4) enhanced cell resistance to oxidative and thermal stresses; and 5) accelerated degradation of neutral lipids deposited in lipid droplets. Our findings provide new insights into mechanisms through which chemicals extracted from certain plants can slow biological aging. PMID:26918729

21 CFR 172.898 - Bakers yeast glycan.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Bakers yeast glycan. 172.898 Section 172.898 Food... Bakers yeast glycan. Bakers yeast glycan may be safely used in food in accordance with the following conditions: (a) Bakers yeast glycan is the comminuted, washed, pasteurized, and dried cell walls of the yeast...

Nutrient-regulated Phosphorylation of ATG13 Inhibits Starvation-induced Autophagy*

PubMed Central

Puente, Cindy; Hendrickson, Ronald C.; Jiang, Xuejun

2016-01-01

Autophagy is a conserved catabolic process that utilizes a defined series of membrane trafficking events to generate a de novo double-membrane vesicle termed the autophagosome, which matures by fusing to the lysosome. Subsequently, the lysosome facilitates the degradation and recycling of the cytoplasmic cargo. In yeast, the upstream signals that regulate the induction of starvation-induced autophagy are clearly defined. The nutrient-sensing kinase Tor inhibits the activation of autophagy by regulating the formation of the Atg1-Atg13-Atg17 complex, through hyperphosphorylation of Atg13. However, in mammals, the ortholog complex ULK1-ATG13-FIP200 is constitutively formed. As such, the molecular mechanism by which mTOR regulates mammalian autophagy is unknown. Here we report the identification and characterization of novel nutrient-regulated phosphorylation sites on ATG13: Ser-224 and Ser-258. mTOR directly phosphorylates ATG13 on Ser-258 while Ser-224 is modulated by the AMPK pathway. In ATG13 knock-out cells reconstituted with an unphosphorylatable mutant of ATG13, ULK1 kinase activity is more potent, and amino acid starvation induced more rapid ATG13 and ULK1 translocation. These events culminated in a more rapid starvation-induced autophagy response. Therefore, ATG13 phosphorylation plays a crucial role in autophagy regulation. PMID:26801615

DNA Damage and Genomic Instability Induced by Inappropriate DNA Re-replication

DTIC Science & Technology

2006-04-01

replication in yeast cells. In the prior reporting period we demonstrated that re-replication induces a rapid and significant decrease in cell viability...repair, DNA replication, checkpoint, cell cycle, yeast , RAD9 16. SECURITY CLASSIFICATION OF: 18. NUMBER OF PAGES 19a. NAME OF RESPONSIBLE PERSON...initiation, our laboratory has been able to conditionally induce varying amounts of re- replication in yeast cells. Effectively, cells enter, but do not

Capillary electrophoresis with laser-induced fluorescence detection for studying amino acid uptake by yeast during beer fermentation.

PubMed

Turkia, Heidi; Sirén, Heli; Penttilä, Merja; Pitkänen, Juha-Pekka

2015-01-01

The amino acid composition of cultivation broth is known to affect the biomass accumulation, productivity, and vitality of yeast during cultivation. A separation method based on capillary electrophoresis with laser-induced fluorescence (LIF) detection was developed for the determination of amino acid consumption by Saccharomyces cerevisiae during beer fermentation. Intraday relative standard deviations were less than 2.1% for migration times and between 2.9% and 9.9% for peak areas. Interday relative standard deviations were less than 2.5% for migration times and between 4.4% and 18.9% for peak areas. The quantification limit was even as low as 62.5 pM which equals to below attomole level detection. The method was applied to study the rate of amino acid utilization during beer fermentation. Copyright © 2014 Elsevier B.V. All rights reserved.

The Potent Humanin Analogue (HNG) Protects Germ Cells and Leucocytes While Enhancing Chemotherapy-Induced Suppression of Cancer Metastases in Male Mice.

PubMed

Lue, YanHe; Swerdloff, Ronald; Wan, Junxiang; Xiao, Jialin; French, Samuel; Atienza, Vince; Canela, Victor; Bruhn, Kevin W; Stone, Brian; Jia, Yue; Cohen, Pinchas; Wang, Christina

2015-12-01

Humanin is a peptide that is cytoprotective against stresses in many cell types. We investigated whether a potent humanin analogue S14G-humanin (HNG) would protect against chemotherapy-induced damage to normal cells without interfering with the chemotherapy-induced suppression of cancer cells. Young adult male mice were inoculated iv with murine melanoma cells. After 1 week, cancer-bearing mice were randomized to receive either: no treatment, daily ip injection of HNG, a single ip injection of cyclophosphamide (CP), or CP+HNG and killed at the end of 3 weeks. HNG rescued the CP-induced suppression of leucocytes and protected germ cell from CP-induced apoptosis. Lung metastases were suppressed by HNG or CP alone, and further suppressed by CP+HNG treatment. Plasma IGF-1 levels were suppressed by HNG with or without CP treatment. To investigate whether HNG maintains its protective effects on spermatogonial stem cells, sperm output, and peripheral leucocytes after repeated doses of CP, normal adult male mice received: no treatment, daily sc injection of HNG, 6 ip injections of CP at 5-day intervals, and the same regimens of CP+HNG and killed at the end of 4 weeks of treatment. Cauda epididymal sperm counts were elevated by HNG and suppressed by CP. HNG rescued the CP-induced suppression of spermatogonial stem cells, sperm count and peripheral leucocytes. We conclude that HNG 1) protects CP-induced loss of male germ cells and leucocytes, 2) enhances CP-induced suppression of cancer metastases, and 3) acts as a caloric-restriction mimetic by suppressing IGF-1 levels. Our findings suggest that humanin analogues may be promising adjuvants to chemotherapy.

The Potent Humanin Analogue (HNG) Protects Germ Cells and Leucocytes While Enhancing Chemotherapy-Induced Suppression of Cancer Metastases in Male Mice

PubMed Central

Lue, YanHe; Swerdloff, Ronald; Wan, Junxiang; Xiao, Jialin; French, Samuel; Atienza, Vince; Canela, Victor; Bruhn, Kevin W.; Stone, Brian; Jia, Yue; Cohen, Pinchas

2015-01-01

Humanin is a peptide that is cytoprotective against stresses in many cell types. We investigated whether a potent humanin analogue S14G-humanin (HNG) would protect against chemotherapy-induced damage to normal cells without interfering with the chemotherapy-induced suppression of cancer cells. Young adult male mice were inoculated iv with murine melanoma cells. After 1 week, cancer-bearing mice were randomized to receive either: no treatment, daily ip injection of HNG, a single ip injection of cyclophosphamide (CP), or CP+HNG and killed at the end of 3 weeks. HNG rescued the CP-induced suppression of leucocytes and protected germ cell from CP-induced apoptosis. Lung metastases were suppressed by HNG or CP alone, and further suppressed by CP+HNG treatment. Plasma IGF-1 levels were suppressed by HNG with or without CP treatment. To investigate whether HNG maintains its protective effects on spermatogonial stem cells, sperm output, and peripheral leucocytes after repeated doses of CP, normal adult male mice received: no treatment, daily sc injection of HNG, 6 ip injections of CP at 5-day intervals, and the same regimens of CP+HNG and killed at the end of 4 weeks of treatment. Cauda epididymal sperm counts were elevated by HNG and suppressed by CP. HNG rescued the CP-induced suppression of spermatogonial stem cells, sperm count and peripheral leucocytes. We conclude that HNG 1) protects CP-induced loss of male germ cells and leucocytes, 2) enhances CP-induced suppression of cancer metastases, and 3) acts as a caloric-restriction mimetic by suppressing IGF-1 levels. Our findings suggest that humanin analogues may be promising adjuvants to chemotherapy. PMID:26384090

Opportunistic Pathogenic Yeasts

NASA Astrophysics Data System (ADS)

Banerjee, Uma

Advances in medical research, made during the last few decades, have improved the prophylactic, diagnostic and therapeutic capabilities for variety of infections/diseases. However, many of the prophylactic and therapeutic procedures have been seen in many instances to exact a price of host-vulnerability to an expanding group of opportunistic pathogens and yeasts are one of the important members in it. Fortunately amongst the vast majority of yeasts present in nature only few are considered to have the capability to cause infections when certain opportunities predisposes and these are termed as ‘opportunistic pathogenic yeasts.’ However, the term ‘pathogenic’ is quite tricky, as it depends of various factors of the host, the ‘bug’ and the environment to manifest the clinical infection. The borderline is expanding. In the present century with unprecedented increase in number of immune-compromised host in various disciplines of health care settings, where any yeast, which has the capability to grow at 37 ° C (normal body temperature of human), can be pathogenic and cause infection in particular situation

Tanshinones extend chronological lifespan in budding yeast Saccharomyces cerevisiae.

PubMed

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

2014-10-01

Natural products with anti-aging property have drawn great attention recently but examples of such compounds are exceedingly scarce. By applying a high-throughput assay based on yeast chronological lifespan measurement, we screened the anti-aging activity of 144 botanical materials and found that dried roots of Salvia miltiorrhiza Bunge have significant anti-aging activity. Tanshinones isolated from the plant including cryptotanshione, tanshinone I, and tanshinone IIa, are the active components. Among them, cryptotanshinone can greatly extend the budding yeast Saccharomyces cerevisiae chronological lifespan (up to 2.5 times) in a dose- and the-time-of-addition-dependent manner at nanomolar concentrations without disruption of cell growth. We demonstrate that cryptotanshinone prolong chronological lifespan via a nutrient-dependent regime, especially essential amino acid sensing, and three conserved protein kinases Tor1, Sch9, and Gcn2 are required for cryptotanshinone-induced lifespan extension. In addition, cryptotanshinone significantly increases the lifespan of SOD2-deleted mutants. Altogether, those data suggest that cryptotanshinone might be involved in the regulation of, Tor1, Sch9, Gcn2, and Sod2, these highly conserved longevity proteins modulated by nutrients from yeast to humans.

Functional Analysis With a Barcoder Yeast Gene Overexpression System

PubMed Central

Douglas, Alison C.; Smith, Andrew M.; Sharifpoor, Sara; Yan, Zhun; Durbic, Tanja; Heisler, Lawrence E.; Lee, Anna Y.; Ryan, Owen; Göttert, Hendrikje; Surendra, Anu; van Dyk, Dewald; Giaever, Guri; Boone, Charles; Nislow, Corey; Andrews, Brenda J.

2012-01-01

Systematic analysis of gene overexpression phenotypes provides an insight into gene function, enzyme targets, and biological pathways. Here, we describe a novel functional genomics platform that enables a highly parallel and systematic assessment of overexpression phenotypes in pooled cultures. First, we constructed a genome-level collection of ~5100 yeast barcoder strains, each of which carries a unique barcode, enabling pooled fitness assays with a barcode microarray or sequencing readout. Second, we constructed a yeast open reading frame (ORF) galactose-induced overexpression array by generating a genome-wide set of yeast transformants, each of which carries an individual plasmid-born and sequence-verified ORF derived from the Saccharomyces cerevisiae full-length EXpression-ready (FLEX) collection. We combined these collections genetically using synthetic genetic array methodology, generating ~5100 strains, each of which is barcoded and overexpresses a specific ORF, a set we termed “barFLEX.” Additional synthetic genetic array allows the barFLEX collection to be moved into different genetic backgrounds. As a proof-of-principle, we describe the properties of the barFLEX overexpression collection and its application in synthetic dosage lethality studies under different environmental conditions. PMID:23050238

Secretion metabolites of probiotic yeast, Pichia kudriavzevii AS-12, induces apoptosis pathways in human colorectal cancer cell lines.

PubMed

Saber, Amir; Alipour, Beitollah; Faghfoori, Zeinab; Mousavi Jam, Ali; Yari Khosroushahi, Ahmad

2017-05-01

There is a common agreement on the important role of the gastrointestinal microbiota in the etiology of cancer. Benign probiotic yeast strains are able to ameliorate intestinal microbiota and regulate the host metabolism, physiology, and immune system through anti-inflammatory, antiproliferative, and anticancer effects. We hypothesized that Pichia kudriavzevii AS-12 secretion metabolites possess anticancer activity on human colorectal cancer cells (HT-29, Caco-2) via inhibiting growth and inducing apoptosis. This study aimed to assess the anticancer effect of P. kudriavzevii AS-12 secretion metabolites and the underlying mechanisms. The cytotoxicity evaluations were performed via 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide assay; 4',6-diamidino-2-phenylindole staining; and FACS-flow cytometry tests. Also, the effects of P. kudriavzevii AS-12 secretion metabolites on the expression level of 6 important genes (BAD, Bcl-2, Caspase-3, Caspase-8, Caspase-9 and Fas-R) involved in the extrinsic and intrinsic apoptosis pathways were studied by real-time polymerase chain reaction method. P. kudriavzevii AS-12 secretion metabolites showed significant (P < .0001) cytotoxic effects on HT-29 cells (57.5%) and Caco-2 (32.5%) compared to KDR/293 normal cells (25%). Moreover, the cytotoxic effects of examined yeast supernatant on HT-29 cells were comparable with 5-fluorouracil, as a positive control (57.5% versus 62.2% respectively). Flow cytometric results showed that the induction of apoptosis is the main mechanism of the anticancer effects. Also, according to the reverse transcriptase polymerase chain reaction results, the expression level of proapoptotic genes (BAD, Caspase-3, Caspase-8, Caspase-9, and Fas-R) in treated HT-29 and Caco-2 cells was higher than untreated and normal cells, whereas the antiapoptotic gene (Bcl-2) was downregulated. P. kudriavzevii AS-12 secretion metabolites exert its anticancer effects by inhibiting cell proliferation and inducing

Cellular immune responses in patients with hepatitis B surface antigen seroclearance induced by antiviral therapy

PubMed Central

2011-01-01

Background The mechanisms by which chronic hepatitis B is completely resolved through antiviral therapy are unknown, and the contribution of acquired T cell immunity to hepatitis B surface antigen (HBsAg) seroclearance has not been investigated. Therefore, we measured the T-cell responses to core and envelope antigens in patients with HBsAg seroclearance. Methods Fourteen subjects with HBsAg seroclearance following antiviral treatment for chronic hepatitis B, 7 HBeAg-positive immunotolerant HBV carriers and 9 HBeAg-negative inactive HBsAg carriers were recruited. HBV-specific T-cell responses to recombinant HBV core (rHBcAg) and envelope (rHBsAg) proteins and pools of core and envelope peptides were measured using an ELISPOT assay detecting interferon-gamma and intracellular cytokine staining (ICS) assays detecting interferon-gamma or interleukin 2. Results Interferon-gamma ELISPOT assays showed a low frequency of weak responses to the rHBsAg and S peptide pool in the HBsAg seroclearance group, and the response frequency to the rHBcAg and the C peptide pool was higher than to the rHBsAg (P < 0.001) and S peptide pool (P = 0.001) respectively. A higher response frequency to C than S peptide pools was confirmed in the interferon-gamma ICS assays for both CD4+ (P = 0.033) and CD8+ (P = 0.040) T cells in the HBsAg seroclearance group. The responses to C and S antigens in the inactive carriers were similar. Conclusions There was a low frequency of CD4+ and CD8+ T cell immune responses to envelope antigens in Chinese subjects with HBsAg seroclearance following antiviral therapy. It is unlikely that these immune responses are responsible for HBsAg seroclearance in these subjects. PMID:21320337

Genome-wide expression analyses of the stationary phase model of ageing in yeast.

PubMed

Wanichthanarak, Kwanjeera; Wongtosrad, Nutvadee; Petranovic, Dina

2015-07-01

Ageing processes involved in replicative lifespan (RLS) and chronological lifespan (CLS) have been found to be conserved among many organisms, including in unicellular Eukarya such as yeast Saccharomyces cerevisiae. Here we performed an integrated approach of genome wide expression profiles of yeast at different time points, during growth and starvation. The aim of the study was to identify transcriptional changes in those conditions by using several different computational analyses in order to propose transcription factors, biological networks and metabolic pathways that seem to be relevant during the process of chronological ageing in yeast. Specifically, we performed differential gene expression analysis, gene-set enrichment analysis and network-based analysis, and we identified pathways affected in the stationary phase and specific transcription factors driving transcriptional adaptations. The results indicate signal propagation from G protein-coupled receptors through signaling pathway components and other stress and nutrient-induced transcription factors resulting in adaptation of yeast cells to the lack of nutrients by activating metabolism associated with aerobic metabolism of carbon sources such as ethanol, glycerol and fatty acids. In addition, we found STE12, XBP1 and TOS8 as highly connected nodes in the subnetworks of ageing yeast. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Yeast Droplets

NASA Astrophysics Data System (ADS)

Nguyen, Baochi; Upadhyaya, Arpita; van Oudenaarden, Alexander; Brenner, Michael

2002-11-01

It is well known that the Young's law and surface tension govern the shape of liquid droplets on solid surfaces. Here we address through experiments and theory the shape of growing aggregates of yeast on agar substrates, and assess whether these ideas still hold. Experiments are carried out on Baker's yeast, with different levels of expressions of an adhesive protein governing cell-cell and cell-substrate adhesion. Changing either the agar concentration or the expression of this protein modifies the local contact angle of a yeast droplet. When the colony is small, the shape is a spherical cap with the contact angle obeying Young's law. However, above a critical volume this structure is unstable, and the droplet becomes nonspherical. We present a theoretical model where this instability is caused by bulk elastic effects. The model predicts that the transition depends on both volume and contact angle, in a manner quantitatively consistent with our experiments.

Ischemia--reperfusion: a look from yeast mitochondria.

PubMed

Stella, Carlos; Burgos, Isabel; Chapela, S; Gamondi, Oliver

2011-01-01

The apoptotic phenomena observed in tissues which are subdued to ischemia and then to technical therapeutics of perfusion keep causing serious problems in the patient's clinical recovery. Then, they constitute a challenge to resolve. The objective of this work is to discuss the intracellular mechanisms that lead cells to apoptosis during the ischemia-reperfusion process, taking into consideration that these phenomena are observable in a simple microorganism as the yeast Saccharomyces cerevisiae. Yeast provide an alternative study system in which the effects of certain cytoprotective drugs can be evaluated. The results can then be extrapolated to other types of cells. Several works have focused on the role of mitochondria in the apoptotic processes of cellular necrosis. One of the main factors responsible for this process is the unregulated opening of the permeability barrier. The inner membrane thus allows the unrestricted passage of ions and the release of apoptotic mediators from the inner membrane space towards the cytosol. Also, there is an increase in the level of reactive oxygen species (ROS) and the uncoupling of oxidative phosphorylation, which lead to the reversal of ATP synthesis to ATP hydrolysis. The driving cause of this complex process is the opening of an non-specific pore located in the mitochondrial membrane, denominated mammalian permeability transition pore (mPTP), which is also expressed in yeast (yPTP). From the functional point of view, the yeast pore presents some of the characteristics observed in mammals, and is similar in the defensive response against the deleterious mechanisms caused by oxidative stress. An increasing body of evidence supports the concept that the pharmacological inhibition of the mPTP is an actual and promising strategy for the protection of tissues in ischemic situations in order to avoid the damage induced by perfusion.

Electric field mediated loading of macromolecules in intact yeast cells is critically controlled at the wall level.

PubMed

Ganeva, V; Galutzov, B; Teissié, J

1995-12-13

The mechanism of electric field mediated macromolecule transfer inside an intact yeast cell was investigated by observing, under a microscope, the fluorescence associated to cells after pulsation in a buffer containing two different hydrophilic fluorescent dyes. In the case of a small probe such as propidium iodide, a long lived permeabilized state was induced by the field as classically observed on wall free systems. Penetration of a 70 kDa FITC dextran was obtained only by using drastic conditions and only a very limited number of yeast cells which took up macromolecules remained viable. Most dextrans were trapped in the wall. A dramatic improvement in transfer of dextrans was observed when the cells were treated by dithiothreitol before pulsation. A cytoplasmic protein leakage was detected after the electric treatment suggesting that an irreversible damage took place in the walls of many pulsed cells. Electroloading of macromolecules in intact yeast cells appears to be controlled by a field induced short lived alteration of the envelope organization.

Quercetin Protects Yeast Saccharomyces cerevisiae pep4 Mutant from Oxidative and Apoptotic Stress and Extends Chronological Lifespan.

PubMed

Alugoju, Phaniendra; Janardhanshetty, Sudharshan Setra; Subaramanian, Subasri; Periyasamy, Latha; Dyavaiah, Madhu

2018-05-01

The yeast Saccharomyces cerevisiae PEP4 gene encodes vacuolar endopeptidase proteinase A (Pep4p), which is a homolog of the human CTSD gene that encodes cathepsin D. Mutation of CTSD gene in human resulted in a number of neurodegenerative diseases. In this study, we have shown that yeast pep4 mutant cells are highly sensitive to oxidative and apoptotic stress induced by hydrogen peroxide and acetic acid, respectively. pep4∆ cells also showed accumulation of reactive oxygen species (ROS), apoptotic markers, and reduced chronological lifespan. In contrast, quercetin pretreatment protected the pep4 mutant from oxidative and apoptotic stress-induced sensitivity by scavenging ROS and reducing apoptotic markers. The percentage viability of quercetin-treated pep4∆ cells was more pronounced and increased stress resistance against oxidant, apoptotic, and heat stress during chronological aging. From our experimental results, we concluded that quercetin protects yeast pep4 mutant cells from oxidative stress and apoptosis, thereby increasing viability during chronological aging.

Tricyclic Compounds Containing Non-enolizable Cyano Enones. A Novel Class of Highly Potent Anti-inflammatory and Cytoprotective Agents=

PubMed Central

Honda, Tadashi; Yoshizawa, Hidenori; Sundararajan, Chitra; David, Emilie; Lajoie, Marc J.; Favaloro, Frank G.; Janosik, Tomasz; Su, Xiaobo; Honda, Yukiko; Roebuck, Bill D.; Gribble, Gordon W.

2011-01-01

Forty-four novel tricycles containing non-enolizable cyano enones (TCEs) were designed and synthesized on the basis of a semisynthetic pentacyclic triterpenoid, bardoxolone methyl, which is currently being developed in Phase II clinical trials for the treatment of severe chronic kidney disease in diabetic patients. Most of the TCEs having two different kinds of non-enolizable cyano enones in rings A and C are highly potent suppressors of induction of inducible nitric oxide synthase stimulated with interferon-γ, and highly potent inducers of the cytoprotective enzymes heme oxygenase-1 and NAD(P)H:quinone oxidoreductase-1. Among these compounds, (±)-(4bS,8aR,10aS)-10a-ethynyl-4b,8,8-trimethyl-3,7-dioxo-3,4b,7,8,8a,9,10,10a-octahydrophenanthrene-2,6-dicarbonitrile ((±)-31) is the most potent in these bioassays in our pool of drug candidates including semisynthetic triterpenoids and synthetic tricycles. These facts strongly suggest that an essential factor for potency is not a triterpenoid skeleton, but the cyano enone functionality. Notably, TCE 31 reduces hepatic tumorigenesis induced with aflatoxin in rats. Further preclinical studies and detailed mechanism studies on 31 are in progress. PMID:21361338

An Investigation into Rumen Fungal and Protozoal Diversity in Three Rumen Fractions, during High-Fiber or Grain-Induced Sub-Acute Ruminal Acidosis Conditions, with or without Active Dry Yeast Supplementation

PubMed Central

Ishaq, Suzanne L.; AlZahal, Ousama; Walker, Nicola; McBride, Brian

2017-01-01

Sub-acute ruminal acidosis (SARA) is a gastrointestinal functional disorder in livestock characterized by low rumen pH, which reduces rumen function, microbial diversity, host performance, and host immune function. Dietary management is used to prevent SARA, often with yeast supplementation as a pH buffer. Almost nothing is known about the effect of SARA or yeast supplementation on ruminal protozoal and fungal diversity, despite their roles in fiber degradation. Dairy cows were switched from a high-fiber to high-grain diet abruptly to induce SARA, with and without active dry yeast (ADY, Saccharomyces cerevisiae) supplementation, and sampled from the rumen fluid, solids, and epimural fractions to determine microbial diversity using the protozoal 18S rRNA and the fungal ITS1 genes via Illumina MiSeq sequencing. Diet-induced SARA dramatically increased the number and abundance of rare fungal taxa, even in fluid fractions where total reads were very low, and reduced protozoal diversity. SARA selected for more lactic-acid utilizing taxa, and fewer fiber-degrading taxa. ADY treatment increased fungal richness (OTUs) but not diversity (Inverse Simpson, Shannon), but increased protozoal richness and diversity in some fractions. ADY treatment itself significantly (P < 0.05) affected the abundance of numerous fungal genera as seen in the high-fiber diet: Lewia, Neocallimastix, and Phoma were increased, while Alternaria, Candida Orpinomyces, and Piromyces spp. were decreased. Likewise, for protozoa, ADY itself increased Isotricha intestinalis but decreased Entodinium furca spp. Multivariate analyses showed diet type was most significant in driving diversity, followed by yeast treatment, for AMOVA, ANOSIM, and weighted UniFrac. Diet, ADY, and location were all significant factors for fungi (PERMANOVA, P = 0.0001, P = 0.0452, P = 0.0068, Monte Carlo correction, respectively, and location was a significant factor (P = 0.001, Monte Carlo correction) for protozoa. Diet-induced

An Investigation into Rumen Fungal and Protozoal Diversity in Three Rumen Fractions, during High-Fiber or Grain-Induced Sub-Acute Ruminal Acidosis Conditions, with or without Active Dry Yeast Supplementation.

PubMed

Ishaq, Suzanne L; AlZahal, Ousama; Walker, Nicola; McBride, Brian

2017-01-01

Sub-acute ruminal acidosis (SARA) is a gastrointestinal functional disorder in livestock characterized by low rumen pH, which reduces rumen function, microbial diversity, host performance, and host immune function. Dietary management is used to prevent SARA, often with yeast supplementation as a pH buffer. Almost nothing is known about the effect of SARA or yeast supplementation on ruminal protozoal and fungal diversity, despite their roles in fiber degradation. Dairy cows were switched from a high-fiber to high-grain diet abruptly to induce SARA, with and without active dry yeast (ADY, Saccharomyces cerevisiae ) supplementation, and sampled from the rumen fluid, solids, and epimural fractions to determine microbial diversity using the protozoal 18S rRNA and the fungal ITS1 genes via Illumina MiSeq sequencing. Diet-induced SARA dramatically increased the number and abundance of rare fungal taxa, even in fluid fractions where total reads were very low, and reduced protozoal diversity. SARA selected for more lactic-acid utilizing taxa, and fewer fiber-degrading taxa. ADY treatment increased fungal richness (OTUs) but not diversity (Inverse Simpson, Shannon), but increased protozoal richness and diversity in some fractions. ADY treatment itself significantly ( P < 0.05) affected the abundance of numerous fungal genera as seen in the high-fiber diet: Lewia, Neocallimastix , and Phoma were increased, while Alternaria, Candida Orpinomyces , and Piromyces spp. were decreased. Likewise, for protozoa, ADY itself increased Isotricha intestinalis but decreased Entodinium furca spp. Multivariate analyses showed diet type was most significant in driving diversity, followed by yeast treatment, for AMOVA, ANOSIM, and weighted UniFrac. Diet, ADY, and location were all significant factors for fungi (PERMANOVA, P = 0.0001, P = 0.0452, P = 0.0068, Monte Carlo correction, respectively, and location was a significant factor ( P = 0.001, Monte Carlo correction) for protozoa. Diet-induced

The Antarctic yeast Candida sake: Understanding cold metabolism impact on wine.

PubMed

Ballester-Tomás, Lidia; Prieto, Jose A; Gil, Jose V; Baeza, Marcelo; Randez-Gil, Francisca

2017-03-20

Current winemaking trends include low-temperature fermentations and using non-Saccharomyces yeasts as the most promising tools to produce lower alcohol and increased aromatic complexity wines. Here we explored the oenological attributes of a C. sake strain, H14Cs, isolated in the sub-Antarctic region. As expected, the cold sea water yeast strain showed greater cold growth, Na + -toxicity resistance and freeze tolerance than the S. cerevisiae QA23 strain, which we used as a commercial wine yeast control. C. sake H14Cs was found to be more sensitive to ethanol. The fermentation trials of low-sugar content must demonstrated that C. sake H14Cs allowed the cold-induced lag phase of growth to be eliminated and also notably reduced the ethanol (-30%) and glycerol (-50%) content in wine. Instead C. sake produced sorbitol as a compatible osmolyte. Finally, the inspection of the main wine volatile compounds revealed that C. sake produced more higher alcohols than S. cerevisiae. In conclusion, our work evidences that using the Antarctic C. sake H14Cs yeast improves low-temperature must fermentations and has the potential to provide a wine with less ethanol and also particular attributes. Copyright © 2017 Elsevier B.V. All rights reserved.

Lager Yeast Comes of Age

PubMed Central

2014-01-01

Alcoholic fermentations have accompanied human civilizations throughout our history. Lager yeasts have a several-century-long tradition of providing fresh beer with clean taste. The yeast strains used for lager beer fermentation have long been recognized as hybrids between two Saccharomyces species. We summarize the initial findings on this hybrid nature, the genomics/transcriptomics of lager yeasts, and established targets of strain improvements. Next-generation sequencing has provided fast access to yeast genomes. Its use in population genomics has uncovered many more hybridization events within Saccharomyces species, so that lager yeast hybrids are no longer the exception from the rule. These findings have led us to propose network evolution within Saccharomyces species. This “web of life” recognizes the ability of closely related species to exchange DNA and thus drain from a combined gene pool rather than be limited to a gene pool restricted by speciation. Within the domesticated lager yeasts, two groups, the Saaz and Frohberg groups, can be distinguished based on fermentation characteristics. Recent evidence suggests that these groups share an evolutionary history. We thus propose to refer to the Saaz group as Saccharomyces carlsbergensis and to the Frohberg group as Saccharomyces pastorianus based on their distinct genomes. New insight into the hybrid nature of lager yeast will provide novel directions for future strain improvement. PMID:25084862

Yeast killer systems.

PubMed Central

Magliani, W; Conti, S; Gerloni, M; Bertolotti, D; Polonelli, L

1997-01-01

The killer phenomenon in yeasts has been revealed to be a multicentric model for molecular biologists, virologists, phytopathologists, epidemiologists, industrial and medical microbiologists, mycologists, and pharmacologists. The surprisingly widespread occurrence of the killer phenomenon among taxonomically unrelated microorganisms, including prokaryotic and eukaryotic pathogens, has engendered a new interest in its biological significance as well as its theoretical and practical applications. The search for therapeutic opportunities by using yeast killer systems has conceptually opened new avenues for the prevention and control of life-threatening fungal diseases through the idiotypic network that is apparently exploited by the immune system in the course of natural infections. In this review, the biology, ecology, epidemiology, therapeutics, serology, and idiotypy of yeast killer systems are discussed. PMID:9227858

Calcium-dependent nonspecific permeability of the inner mitochondrial membrane is not induced in mitochondria of the yeast Endomyces magnusii.

PubMed

Deryabina, Y I; Isakova, E P; Shurubor, E I; Zvyagilskaya, R A

2004-09-01

Mitochondria of the yeast Endomyces magnusii were examined for the presence of a Ca2+- and phosphate-induced permeability of the inner mitochondrial membrane (pore). For this purpose, coupled mitochondria were incubated under conditions known to induce the permeability transition pore in animal mitochondria, i.e., in the presence of high concentrations of Ca2+ and P(i), prooxidants (t-butylhydroperoxide), oxaloacetate, atractyloside (an inhibitor of ADP/ATP translocator), SH-reagents, by depletion of adenine nucleotide pools, and deenergization of the mitochondria. Large amplitude swelling, collapse of the membrane potential, and efflux of the accumulated Ca2+ were used as parameters for demonstrating pore induction. E. magnusii mitochondria were highly resistant to the above-mentioned substances. Deenergization of mitochondria or depletion of adenine nucleotide pools have no effect on low-amplitude swelling or the other parameters. Cyclosporin A, a specific inhibitor of the nonspecific permeability transition in animal mitochondria, did not affect the parameters measured. It is thus evident that E. magnusii mitochondria lack a functional Ca2+-dependent pore, or possess a pore differently regulated as compared to that of mammalian mitochondria.

Heat Shock Partially Dissociates the Overlapping Modules of the Yeast Protein-Protein Interaction Network: A Systems Level Model of Adaptation

PubMed Central

Mihalik, Ágoston; Csermely, Peter

2011-01-01

Network analysis became a powerful tool giving new insights to the understanding of cellular behavior. Heat shock, the archetype of stress responses, is a well-characterized and simple model of cellular dynamics. S. cerevisiae is an appropriate model organism, since both its protein-protein interaction network (interactome) and stress response at the gene expression level have been well characterized. However, the analysis of the reorganization of the yeast interactome during stress has not been investigated yet. We calculated the changes of the interaction-weights of the yeast interactome from the changes of mRNA expression levels upon heat shock. The major finding of our study is that heat shock induced a significant decrease in both the overlaps and connections of yeast interactome modules. In agreement with this the weighted diameter of the yeast interactome had a 4.9-fold increase in heat shock. Several key proteins of the heat shock response became centers of heat shock-induced local communities, as well as bridges providing a residual connection of modules after heat shock. The observed changes resemble to a ‘stratus-cumulus’ type transition of the interactome structure, since the unstressed yeast interactome had a globally connected organization, similar to that of stratus clouds, whereas the heat shocked interactome had a multifocal organization, similar to that of cumulus clouds. Our results showed that heat shock induces a partial disintegration of the global organization of the yeast interactome. This change may be rather general occurring in many types of stresses. Moreover, other complex systems, such as single proteins, social networks and ecosystems may also decrease their inter-modular links, thus develop more compact modules, and display a partial disintegration of their global structure in the initial phase of crisis. Thus, our work may provide a model of a general, system-level adaptation mechanism to environmental changes. PMID:22022244

Yeasts as distinct life forms of fungi

USDA-ARS?s Scientific Manuscript database

This review describes all presently recognized genera of the Ascomycete yeasts (Saccharomycotina, budding yeasts, and the Taphrinomycotina, fission yeasts and related) as well as all currently recognized genera of the Basidiomycete yeasts. This update will be the lead chapter for a book entitled “Ye...

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

Evolutionary History of Ascomyceteous Yeasts

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

Haridas, Sajeet; Riley, Robert; Salamov, Asaf

2014-06-06

Yeasts are important for many industrial and biotechnological processes and show remarkable diversity despite morphological similarities. We have sequenced the genomes of 16 ascomycete yeasts of taxonomic and industrial importance including members of Saccharomycotina and Taphrinomycotina. A comparison of these with several other previously published yeast genomes have added increased confidence to the phylogenetic positions of previously poorly placed species including Saitoella complicata, Babjeviella inositovora and Metschnikowia bicuspidata. Phylogenetic analysis also showed that yeasts with alternative nuclear codon usage where CUG encodes serine instead of leucine are monophyletic within the Saccharomycotina. Most of the yeasts have compact genomes with amore » large fraction of single exon genes with Lipomyces starkeyi and the previously published Pneumocystis jirovecii being notable exceptions. Intron analysis suggests that early diverging species have more introns. We also observed a large number of unclassified lineage specific non-simple repeats in these genomes.« less

The potent antimicrobial properties of cell penetrating peptide-conjugated silver nanoparticles with excellent selectivity for gram-positive bacteria over erythrocytes.

PubMed

Liu, Lihong; Yang, Jun; Xie, Jianping; Luo, Zhentao; Jiang, Jiang; Yang, Yi Yan; Liu, Shaomin

2013-05-07

Silver nanoparticles are of great interest for use as antimicrobial agents. Studies aimed at producing potent nano-silver biocides have focused on manipulation of particle size, shape, composition and surface charge. Here, we report the cell penetrating peptide catalyzed formation of antimicrobial silver nanoparticles in N,N-dimethylformamide. The novel nano-composite demonstrated a distinctly enhanced biocidal effect toward bacteria (gram-positive Bacillus subtilis, gram-negative Escherichia coli) and pathogenic yeast (Candida albicans), as compared to triangular and extremely small silver nanoparticles. In addition, a satisfactory biocompatibility was verified by a haemolysis test. Our results provide a paradigm in developing strategies that can maximize the silver nanoparticle application potentials while minimizing the toxic effects.

The yeast Hsp70 Ssa1 is a sensor for activation of the heat shock response by thiol-reactive compounds

PubMed Central

Wang, Yanyu; Gibney, Patrick A.; West, James D.; Morano, Kevin A.

2012-01-01

The heat shock transcription factor HSF1 governs the response to heat shock, oxidative stresses, and xenobiotics through unknown mechanisms. We demonstrate that diverse thiol-reactive molecules potently activate budding yeast Hsf1. Hsf1 activation by thiol-reactive compounds is not consistent with the stresses of misfolding of cytoplasmic proteins or cytotoxicity. Instead, we demonstrate that the Hsp70 chaperone Ssa1, which represses Hsf1 in the absence of stress, is hypersensitive to modification by a thiol-reactive probe. Strikingly, mutation of two conserved cysteine residues to serine in Ssa1 rendered cells insensitive to Hsf1 activation and subsequently induced thermotolerance by thiol-reactive compounds, but not by heat shock. Conversely, substitution with the sulfinic acid mimic aspartic acid resulted in constitutive Hsf1 activation. Cysteine 303, located within the nucleotide-binding domain, was found to be modified in vivo by a model organic electrophile, demonstrating that Ssa1 is a direct target for thiol-reactive molecules through adduct formation. These findings demonstrate that Hsp70 is a proximal sensor for Hsf1-mediated cytoprotection and can discriminate between two distinct environmental stressors. PMID:22809627

Characterization of karyopherins in androgen receptor intracellular trafficking in the yeast model

PubMed Central

Nguyen, Minh M; Harmon, Robert M; Wang, Zhou

2014-01-01

Background: Mechanisms regulating androgen receptor (AR) subcellular localization represent an essential component of AR signaling. Karyopherins are a family of nucleocytoplasmic trafficking factors. In this paper, we used the yeast model to study the effects of karyopherins on the subcellular localization of the AR. Methods: Yeast mutants deficient in different nuclear transport factors were transformed with various AR based, GFP tagged constructs and their localization was monitored using microscopy. Results: We showed that yeast can mediate androgen-induced AR nuclear localization and that in addition to the import factor, Importinα/β, this process required the import karyopherin Sxm1. We also showed that a previously identified nuclear export sequence (NESAR) in the ligand binding domain of AR does not appear to rely on karyopherins for cytoplasmic localization. Conclusions: These results suggest that while AR nuclear import relies on karyopherin activity, AR nuclear export and/or cytoplasmic localization may require other undefined mechanisms. PMID:25031696

A common bacterial metabolite elicits prion-based bypass of glucose repression

PubMed Central

Garcia, David M; Dietrich, David; Clardy, Jon; Jarosz, Daniel F

2016-01-01

Robust preference for fermentative glucose metabolism has motivated domestication of the budding yeast Saccharomyces cerevisiae. This program can be circumvented by a protein-based genetic element, the [GAR+] prion, permitting simultaneous metabolism of glucose and other carbon sources. Diverse bacteria can elicit yeast cells to acquire [GAR+], although the molecular details of this interaction remain unknown. Here we identify the common bacterial metabolite lactic acid as a strong [GAR+] inducer. Transient exposure to lactic acid caused yeast cells to heritably circumvent glucose repression. This trait had the defining genetic properties of [GAR+], and did not require utilization of lactic acid as a carbon source. Lactic acid also induced [GAR+]-like epigenetic states in fungi that diverged from S. cerevisiae ~200 million years ago, and in which glucose repression evolved independently. To our knowledge, this is the first study to uncover a bacterial metabolite with the capacity to potently induce a prion. DOI: http://dx.doi.org/10.7554/eLife.17978.001 PMID:27906649

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

Biomedical applications of yeast- a patent view, part one: yeasts as workhorses for the production of therapeutics and vaccines.

PubMed

Roohvand, Farzin; Shokri, Mehdi; Abdollahpour-Alitappeh, Meghdad; Ehsani, Parastoo

2017-08-01

Yeasts, as Eukaryotes, offer unique features for ease of growth and genetic manipulation possibilities, making it an exceptional microbial host. Areas covered: This review provides general and patent-oriented insights into production of biopharmaceuticals by yeasts. Patents, wherever possible, were correlated to the original or review articles. The review describes applications of major GRAS (generally regarded as safe) yeasts for the production of therapeutic proteins and subunit vaccines; additionally, immunomodulatory properties of yeast cell wall components were reviewed for use of whole yeast cells as a new vaccine platform. The second part of the review will discuss yeast- humanization strategies and innovative applications. Expert opinion: Biomedical applications of yeasts were initiated by utilization of Saccharomyces cerevisiae, for production of leavened (fermented) products, and advanced to serve to produce biopharmaceuticals. Higher biomass production and expression/secretion yields, more similarity of glycosylation patterns to mammals and possibility of host-improvement strategies through application of synthetic biology might enhance selection of Pichia pastoris (instead of S. cerevisiae) as a host for production of biopharmaceutical in future. Immunomodulatory properties of yeast cell wall β-glucans and possibility of intracellular expression of heterologous pathogen/tumor antigens in yeast cells have expanded their application as a new platform, 'Whole Yeast Vaccines'.

Building Better Biosensors for Exploration into Deep-Space, Using Humanized Yeast

NASA Technical Reports Server (NTRS)

Liddell, Lauren; Santa Maria, Sergio; Tieze, Sofia; Bhattacharya, Sharmila

2017-01-01

1.BioSentinel is 1 of 13 secondary payloads hitching a ride beyond Low Earth Orbit on Exploration Mission 1 (EM-1), set to launch from NASAs Space Launch System in 2019. EM-1 is our first opportunity to investigate the effects of the deep space environment on a eukaryotic biological system, the budding yeast S. cerevisiae. Though separated by a billion years of evolution we share hundreds of genes important for basic cell function, including responses to DNA damage. Thus, yeast is an ideal biosensor for detecting typesextent of damage induced by deep-space radiation.We will fly desiccated cells, then rehydrate to wake them up when the automated payload is ready to initiate the experiment. Rehydration solution contains SC (Synthetic Complete) media and alamarBlue, an indicator for changes in growth and metabolism. Telemetry of LED readings will then allow us to detect how cells respond throughout the mission. The desiccation-rehydration process can be extremely damaging to cells, and can severely diminish our ability to accurately measure and model cellular responses to deep-space radiation. The aim of this study is to develop a better biosensor: yeast strains that are more resistant to desiccation stress. We will over-express known cellular protectants, including hydrophilin Sip18, the protein disaggregase Hsp104, and thioredoxin Trx2, a responder to oxidative stress, then measure cell viability after desiccation to determine which factors improve stress tolerance. Over-expression of SIP18 in wine yeast starter cultures was previously reported to increase viability following desiccation stress by up to 70. Thus, we expect similar improvements in our space-yeast strains. By designing better yeast biosensors we can better prepare for and mitigate the potential dangers of deep-space radiation for future missions.This work is funded by NASAs AES program.

Detection of HbsAg and hATIII genetically modified goats (Caprahircus) by loop-mediated isothermal amplification.

PubMed

Tao, Chenyu; Zhang, Qingde; Zhai, Shanli; Liu, Bang

2013-11-01

In this study, sensitive and rapid detection systems were designed using a loop-mediated isothermal amplification (LAMP) method to detect the genetically modified goats. A set of 4 primers were designed for each exogenous nucleic acids HBsAg and hATIII. The DNA samples were first amplified with the outer and inner primers and released a single-stranded DNA,of which both ends were stem-loop structure. Then one inner primer hybridized with the loop, and initiated displacement synthesis in less than 1 h. The result could be visualized by both agarose gel electrophoresis and unaided eyes directly after adding SYBR GREEN 1. The detection limit of LAMP was ten copies of target molecules, indicating that LAMP was tenfold more sensitive than the classical PCR. Furthermore, all the samples of genetically modified goats were tested positively by LAMP, and the results demonstrated that the LAMP was a rapid and sensitive method for detecting the genetically modified organism.

Hibiscus vitifolius (Linn.) root extracts shows potent protective action against anti-tubercular drug induced hepatotoxicity.

PubMed

Samuel, Anbu Jeba Sunilson John; Mohan, Syam; Chellappan, Dinesh Kumar; Kalusalingam, Anandarajagopal; Ariamuthu, Saraswathi

2012-05-07

The roots of Hibiscus vitifolius Linn. (Malvaceae) is used for the treatment of jaundice in the folklore system of medicine in India. This study is an attempt to evaluate the hepatoprotective activity of the roots of Hibiscus vitifolius against anti-tubercular drug induced hepatotoxicity. Hepatotoxicity was induced in albino rats of either sex by oral administration of a combination of three anti-tubercular drugs. Petroleum ether, chloroform, methanol and aqueous extracts of roots of Hibiscus vitifolius (400mg/kg/day) were evaluated for their possible hepatoprotective potential. All the extracts were found to be safe up to a dose of 2000mg/kg. Among the four extracts studied, oral administration of methanol extract of Hibiscus vitifolius at 400mg/kg showed significant difference in all the parameters when compared to control. There was a significant (P<0.001) reduction in the levels of serum aspartate amino transaminase, alanine amino transferase, alkaline phosphatase, lactate dehydrogenase, total and direct bilirubin, where as an increase was found in the levels of total cholesterol, total protein and albumin. Liver homogenate studies showed a significant increase in the levels of total protein, phospholipids and glycogen, and a reduction in the levels of total lipids, triglycerides, and cholesterol against control animals. In the tissue anti-oxidant studies, we found a significant increase in the levels of catalase and superoxide dismutase, whereas there was marked reduction in the levels of thiobarbituric acid reactive substances, as compared to control. Histology of liver sections of the animals treated with the extracts showed significant reduction of necrosis and fatty formation when compared with control specimens. These findings suggest that the root extracts of Hibiscus vitifolius have potent hepatoprotective activity, thereby justifying its ethnopharmacological claim. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Emergence of DNA Polymerase ε Antimutators That Escape Error-Induced Extinction in Yeast

PubMed Central

Williams, Lindsey N.; Herr, Alan J.; Preston, Bradley D.

2013-01-01

DNA polymerases (Pols) ε and δ perform the bulk of yeast leading- and lagging-strand DNA synthesis. Both Pols possess intrinsic proofreading exonucleases that edit errors during polymerization. Rare errors that elude proofreading are extended into duplex DNA and excised by the mismatch repair (MMR) system. Strains that lack Pol proofreading or MMR exhibit a 10- to 100-fold increase in spontaneous mutation rate (mutator phenotype), and inactivation of both Pol δ proofreading (pol3-01) and MMR is lethal due to replication error-induced extinction (EEX). It is unclear whether a similar synthetic lethal relationship exists between defects in Pol ε proofreading (pol2-4) and MMR. Using a plasmid-shuffling strategy in haploid Saccharomyces cerevisiae, we observed synthetic lethality of pol2-4 with alleles that completely abrogate MMR (msh2Δ, mlh1Δ, msh3Δ msh6Δ, or pms1Δ mlh3Δ) but not with partial MMR loss (msh3Δ, msh6Δ, pms1Δ, or mlh3Δ), indicating that high levels of unrepaired Pol ε errors drive extinction. However, variants that escape this error-induced extinction (eex mutants) frequently emerged. Five percent of pol2-4 msh2Δ eex mutants encoded second-site changes in Pol ε that reduced the pol2-4 mutator phenotype between 3- and 23-fold. The remaining eex alleles were extragenic to pol2-4. The locations of antimutator amino-acid changes in Pol ε and their effects on mutation spectra suggest multiple mechanisms of mutator suppression. Our data indicate that unrepaired leading- and lagging-strand polymerase errors drive extinction within a few cell divisions and suggest that there are polymerase-specific pathways of mutator suppression. The prevalence of suppressors extragenic to the Pol ε gene suggests that factors in addition to proofreading and MMR influence leading-strand DNA replication fidelity. PMID:23307893

Yeast Model Uncovers Dual Roles of Mitochondria in the Action of Artemisinin

PubMed Central

Li, Wei; Mo, Weike; Shen, Dan; Sun, Libo; Wang, Juan; Lu, Shan; Gitschier, Jane M; Zhou, Bing

2005-01-01

Artemisinins, derived from the wormwood herb Artemisia annua, are the most potent antimalarial drugs currently available. Despite extensive research, the exact mode of action of artemisinins has not been established. Here we use yeast, Saccharamyces cerevisiae, to probe the core working mechanism of this class of antimalarial agents. We demonstrate that artemisinin's inhibitory effect is mediated by disrupting the normal function of mitochondria through depolarizing their membrane potential. Moreover, in a genetic study, we identify the electron transport chain as an important player in artemisinin's action: Deletion of NDE1 or NDI1, which encode mitochondrial NADH dehydrogenases, confers resistance to artemisinin, whereas overexpression of NDE1 or NDI1 dramatically increases sensitivity to artemisinin. Mutations or environmental conditions that affect electron transport also alter host's sensitivity to artemisinin. Sensitivity is partially restored when the Plasmodium falciparum NDI1 ortholog is expressed in yeast ndi1 strain. Finally, we showed that artemisinin's inhibitory effect is mediated by reactive oxygen species. Our results demonstrate that artemisinin's effect is primarily mediated through disruption of membrane potential by its interaction with the electron transport chain, resulting in dysfunctional mitochondria. We propose a dual role of mitochondria played during the action of artemisinin: the electron transport chain stimulates artemisinin's effect, most likely by activating it, and the mitochondria are subsequently damaged by the locally generated free radicals. PMID:16170412

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.

The central domain of yeast transcription factor Rpn4 facilitates degradation of reporter protein in human cells.

PubMed

Morozov, A V; Spasskaya, D S; Karpov, D S; Karpov, V L

2014-10-16

Despite high interest in the cellular degradation machinery and protein degradation signals (degrons), few degrons with universal activity along species have been identified. It has been shown that fusion of a target protein with a degradation signal from mammalian ornithine decarboxylase (ODC) induces fast proteasomal degradation of the chimera in both mammalian and yeast cells. However, no degrons from yeast-encoded proteins capable to function in mammalian cells were identified so far. Here, we demonstrate that the yeast transcription factor Rpn4 undergoes fast proteasomal degradation and its central domain can destabilize green fluorescent protein and Alpha-fetoprotein in human HEK 293T cells. Furthermore, we confirm the activity of this degron in yeast. Thus, the Rpn4 central domain is an effective interspecies degradation signal. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Purification and characterisation of a new hypothalamic satiety peptide, cocaine and amphetamine regulated transcript (CART), produced in yeast.

PubMed

Thim, L; Nielsen, P F; Judge, M E; Andersen, A S; Diers, I; Egel-Mitani, M; Hastrup, S

1998-05-29

Cocaine and amphetamine regulated transcript (CART) is a newly discovered hypothalamic peptide with a potent appetite suppressing activity following intracerebroventricular administration. When the mature rat CART sequence encoding CART(1-102) was inserted in the yeast expression plasmid three CART peptides could be purified from the fermentation broth reflecting processing at dibasic sequences. None of these corresponded to the naturally occurring CART(55-102). In order to obtain CART(55-102) the precursor Glu-Glu-Ile-Asp-CART(55-102) has been produced and CART(55-102) was generated by digestion of the precursor with dipeptidylaminopeptidase-1. All four generated CART peptides have been characterised by N-terminal amino acid sequencing and mass spectrometry. The CART peptides contain six cysteine residues and using the yeast expressed CART(62-102) the disulphide bond configuration was found to be I-III, II-V and IV-VI. When the four CART peptides were intracerebroventricularly injected in fasted mice (0.1 to 2.0 microg) they all produced a dose dependent inhibition of food intake.

21 CFR 184.1983 - Bakers yeast extract.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Bakers yeast extract. 184.1983 Section 184.1983... GRAS § 184.1983 Bakers yeast extract. (a) Bakers yeast extract is the food ingredient resulting from concentration of the solubles of mechanically ruptured cells of a selected strain of yeast, Saccharomyces...

21 CFR 184.1983 - Bakers yeast extract.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Bakers yeast extract. 184.1983 Section 184.1983... Listing of Specific Substances Affirmed as GRAS § 184.1983 Bakers yeast extract. (a) Bakers yeast extract... a selected strain of yeast, Saccharomyces cerevisiae. It may be concentrated or dried. (b) The...

21 CFR 184.1983 - Bakers yeast extract.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Bakers yeast extract. 184.1983 Section 184.1983... Listing of Specific Substances Affirmed as GRAS § 184.1983 Bakers yeast extract. (a) Bakers yeast extract... a selected strain of yeast, Saccharomyces cerevisiae. It may be concentrated or dried. (b) The...

Simultaneous inhibition of multiple oncogenic miRNAs by a multi-potent microRNA sponge.

PubMed

Jung, Jaeyun; Yeom, Chanjoo; Choi, Yeon-Sook; Kim, Sinae; Lee, EunJi; Park, Min Ji; Kang, Sang Wook; Kim, Sung Bae; Chang, Suhwan

2015-08-21

The roles of oncogenic miRNAs are widely recognized in many cancers. Inhibition of single miRNA using antagomiR can efficiently knock-down a specific miRNA. However, the effect is transient and often results in subtle phenotype, as there are other miRNAs contribute to tumorigenesis. Here we report a multi-potent miRNA sponge inhibiting multiple miRNAs simultaneously. As a model system, we targeted miR-21, miR-155 and miR-221/222, known as oncogenic miRNAs in multiple tumors including breast and pancreatic cancers. To achieve efficient knockdown, we generated perfect and bulged-matched miRNA binding sites (MBS) and introduced multiple copies of MBS, ranging from one to five, in the multi-potent miRNA sponge. Luciferase reporter assay showed the multi-potent miRNA sponge efficiently inhibited 4 miRNAs in breast and pancreatic cancer cells. Furthermore, a stable and inducible version of the multi-potent miRNA sponge cell line showed the miRNA sponge efficiently reduces the level of 4 target miRNAs and increase target protein level of these oncogenic miRNAs. Finally, we showed the miRNA sponge sensitize cells to cancer drug and attenuate cell migratory activity. Altogether, our study demonstrates the multi-potent miRNA sponge is a useful tool to examine the functional impact of simultaneous inhibition of multiple miRNAs and proposes a therapeutic potential.

Insulin-induced Gene Protein (INSIG)-dependent Sterol Regulation of Hmg2 Endoplasmic Reticulum-associated Degradation (ERAD) in Yeast*

PubMed Central

Theesfeld, Chandra L.; Hampton, Randolph Y.

2013-01-01

Insulin-induced gene proteins (INSIGs) function in control of cellular cholesterol. Mammalian INSIGs exert control by directly interacting with proteins containing sterol-sensing domains (SSDs) when sterol levels are elevated. Mammalian 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase (HMGR) undergoes sterol-dependent, endoplasmic-reticulum (ER)-associated degradation (ERAD) that is mediated by INSIG interaction with the HMGR SSD. The yeast HMGR isozyme Hmg2 also undergoes feedback-regulated ERAD in response to the early pathway-derived isoprene gernanylgeranyl pyrophosphate (GGPP). Hmg2 has an SSD, and its degradation is controlled by the INSIG homologue Nsg1. However, yeast Nsg1 promotes Hmg2 stabilization by inhibiting GGPP-stimulated ERAD. We have proposed that the seemingly disparate INSIG functions can be unified by viewing INSIGs as sterol-dependent chaperones of SSD clients. Accordingly, we tested the role of sterols in the Nsg1 regulation of Hmg2. We found that both Nsg1-mediated stabilization of Hmg2 and the Nsg1-Hmg2 interaction required the early sterol lanosterol. Lowering lanosterol in the cell allowed GGPP-stimulated Hmg2 ERAD. Thus, Hmg2-regulated degradation is controlled by a two-signal logic; GGPP promotes degradation, and lanosterol inhibits degradation. These data reveal that the sterol dependence of INSIG-client interaction has been preserved for over 1 billion years. We propose that the INSIGs are a class of sterol-dependent chaperones that bind to SSD clients, thus harnessing ER quality control in the homeostasis of sterols. PMID:23306196

New fluoroprostaglandin F(2alpha) derivatives with prostanoid FP-receptor agonistic activity as potent ocular-hypotensive agents.

PubMed

Nakajima, Tadashi; Matsugi, Takeshi; Goto, Wakana; Kageyama, Masaaki; Mori, Nobuaki; Matsumura, Yasushi; Hara, Hideaki

2003-12-01

To find new prostanoid FP-receptor agonists possessing potent ocular-hypotensive effects with minimal side effects, we evaluated the agonistic activities of newly synthesized prostaglandin F(2alpha) derivatives for the prostanoid FP-receptor both in vitro and in vivo. The iris constrictions induced by the derivatives and their effects on melanin content were examined using cat isolated iris sphincters and cultured B16 melanoma cells, respectively. The effects of derivative ester forms on miosis and intraocular pressure (IOP) were evaluated in cats and cynomolgus monkeys, respectively. Of these derivatives, 6 out of 12 compounds were more potent iris constrictors, with EC(50) values of 0.6 to 9.4 nM, than a carboxylic acid of latanoprost (EC(50)=13.6 nM). A carboxylic acid of latanoprost (100 microM) significantly increased the melanin content of cultured B16 melanoma cells, but some 15,15-difluoro derivatives, such as AFP-157 and AFP-172, did not. Topically applied AFP-168, AFP-169 and AFP-175 (isopropyl ester, methyl ester and ethyl ester forms, respectively, of AFP-172) induced miosis in cats more potently than latanoprost. AFP-168 (0.0005%) reduced IOP to the same extent as 0.005% latanoprost (for at least 8 h). These findings indicate that 15,15-difluoroprostaglandin F(2alpha) derivatives, especially AFP-168, have more potent prostanoid FP-receptor agonistic activities than latanoprost. Hence, AFP-168 may be worthy of further evaluation as an ocular-hypotensive agent.

Continuous-flow separation of live and dead yeasts using reservoir-based dielectrophoresis (rDEP)

NASA Astrophysics Data System (ADS)

Patel, Saurin; Showers, Daniel; Vedantam, Pallavi; Tzeng, Tzuen-Rong; Qian, Shizhi; Xuan, Xiangchun

2012-11-01

Separating live and dead cells is critical to the diagnosis of early stage diseases and to the efficacy test of drug screening etc. We develop a novel microfluidic approach to continuous separation of yeast cells by viability inside a reservoir. It exploits the cell dielectrophoresis that is induced by the inherent electric field gradient at the reservoir-microchannel junction to selectively trap dead yeasts and continuously sort them from live ones. We term this approach reservoir-based dielectrophoresis (rDEP). The transporting, focusing, and trapping of live and dead yeast cells at the reservoir-microchannel junction are studied separately by varying the DC-biased AC electric fields. These phenomena can all be reasonably predicted by a 2D numerical model. We find that the AC to DC field ratio for live yeast trapping is higher than that for dead cells because the former experiences a weaker rDEP while having a larger electrokinetic mobility. It is this difference in the AC to DC field ratio that enables the viability-based yeast cell separation. The rDEP approach has unique advantages over existing DEP-based techniques such as the occupation of zero channel space and the elimination of in-channel mechanical or electrical parts. NSF

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.

The yeast p53 functional assay: a new tool for molecular epidemiology. Hopes and facts.

PubMed

Fronza, G; Inga, A; Monti, P; Scott, G; Campomenosi, P; Menichini, P; Ottaggio, L; Viaggi, S; Burns, P A; Gold, B; Abbondandolo, A

2000-04-01

The assumption of molecular epidemiology that carcinogens leave fingerprints has suggested that analysis of the frequency, type, and site of mutations in genes frequently altered in carcinogenesis may provide clues to the identification of the factors contributing to carcinogenesis. In this mini-review, we revise the development, and validation of the yeast-based p53 functional assay as a new tool for molecular epidemiology. We show that this assay has some very interesting virtues but also has some drawbacks. The yeast functional assay can be used to determine highly specific mutation fingerprints in the human p53 cDNA sequence. Discrimination is possible when comparing mutation spectra induced by sufficiently different mutagens. However, we also reported that the same carcinogen may induce distinguishable mutation spectra due to known influencing factors.

Drug Uptake, Lipid Rafts, and Vesicle Trafficking Modulate Resistance to an Anticancer Lysophosphatidylcholine Analogue in Yeast*

PubMed Central

Cuesta-Marbán, Álvaro; Botet, Javier; Czyz, Ola; Cacharro, Luis M.; Gajate, Consuelo; Hornillos, Valentín; Delgado, Javier; Zhang, Hui; Amat-Guerri, Francisco; Acuña, A. Ulises; McMaster, Christopher R.; Revuelta, José Luis; Zaremberg, Vanina; Mollinedo, Faustino

2013-01-01

The ether-phospholipid edelfosine, a prototype antitumor lipid (ATL), kills yeast cells and selectively kills several cancer cell types. To gain insight into its mechanism of action, we performed chemogenomic screens in the Saccharomyces cerevisiae gene-deletion strain collection, identifying edelfosine-resistant mutants. LEM3, AGP2, and DOC1 genes were required for drug uptake. Edelfosine displaced the essential proton pump Pma1p from rafts, inducing its internalization into the vacuole. Additional ATLs, including miltefosine and perifosine, also displaced Pma1p from rafts to the vacuole, suggesting that this process is a major hallmark of ATL cytotoxicity in yeast. Radioactive and synthetic fluorescent edelfosine analogues accumulated in yeast plasma membrane rafts and subsequently the endoplasmic reticulum. Although both edelfosine and Pma1p were initially located at membrane rafts, internalization of the drug toward endoplasmic reticulum and Pma1p to the vacuole followed different routes. Drug internalization was not dependent on endocytosis and was not critical for yeast cytotoxicity. However, mutants affecting endocytosis, vesicle sorting, or trafficking to the vacuole, including the retromer and ESCRT complexes, prevented Pma1p internalization and were edelfosine-resistant. Our data suggest that edelfosine-induced cytotoxicity involves raft reorganization and retromer- and ESCRT-mediated vesicular transport and degradation of essential raft proteins leading to cell death. Cytotoxicity of ATLs is mainly dependent on the changes they induce in plasma membrane raft-located proteins that lead to their internalization and subsequent degradation. Edelfosine toxicity can be circumvented by inactivating genes that then result in the recycling of internalized cell-surface proteins back to the plasma membrane. PMID:23335509

Inventions on baker's yeast strains and specialty ingredients.

PubMed

Gélinas, Pierre

2009-06-01

Baker's yeast is one of the oldest food microbial starters. Between 1927 and 2008, 165 inventions on more than 337 baker's yeast strains were patented. The first generation of patented yeast strains claimed improved biomass yield at the yeast plant, higher gassing power in dough or better survival to drying to prepare active dry baker's yeast. Especially between 1980 and 1995, a major interest was given to strains for multiple bakery applications such as dough with variable sugar content and stored at refrigeration (cold) or freezing temperatures. During the same period, genetically engineered yeast strains became very popular but did not find applications in the baking industry. Since year 2000, patented baker's yeast strains claimed aroma, anti-moulding or nutritive properties to better meet the needs of the baking industry. In addition to patents on yeast strains, 47 patents were issued on baker's yeast specialty ingredients for niche markets. This review shows that patents on baker's yeast with improved characteristics such as aromatic or nutritive properties have regularly been issued since the 1920's. Overall, it also confirms recent interest for a very wide range of tailored-made yeast-based ingredients for bakery applications.

21 CFR 172.898 - Bakers yeast glycan.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Bakers yeast glycan. 172.898 Section 172.898 Food... Multipurpose Additives § 172.898 Bakers yeast glycan. Bakers yeast glycan may be safely used in food in accordance with the following conditions: (a) Bakers yeast glycan is the comminuted, washed, pasteurized, and...

21 CFR 172.898 - Bakers yeast glycan.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Bakers yeast glycan. 172.898 Section 172.898 Food... Multipurpose Additives § 172.898 Bakers yeast glycan. Bakers yeast glycan may be safely used in food in accordance with the following conditions: (a) Bakers yeast glycan is the comminuted, washed, pasteurized, and...

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.

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

Tombusviruses upregulate phospholipid biosynthesis via interaction between p33 replication protein and yeast lipid sensor proteins during virus replication in yeast

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

Barajas, Daniel; Xu, Kai; Sharma, Monika

Positive-stranded RNA viruses induce new membranous structures and promote membrane proliferation in infected cells to facilitate viral replication. In this paper, the authors show that a plant-infecting tombusvirus upregulates transcription of phospholipid biosynthesis genes, such as INO1, OPI3 and CHO1, and increases phospholipid levels in yeast model host. This is accomplished by the viral p33 replication protein, which interacts with Opi1p FFAT domain protein and Scs2p VAP protein. Opi1p and Scs2p are phospholipid sensor proteins and they repress the expression of phospholipid genes. Accordingly, deletion of OPI1 transcription repressor in yeast has a stimulatory effect on TBSV RNA accumulation andmore » enhanced tombusvirus replicase activity in an in vitro assay. Altogether, the presented data convincingly demonstrate that de novo lipid biosynthesis is required for optimal TBSV replication. Overall, this work reveals that a (+)RNA virus reprograms the phospholipid biosynthesis pathway in a unique way to facilitate its replication in yeast cells. - Highlights: • Tombusvirus p33 replication protein interacts with FFAT-domain host protein. • Tombusvirus replication leads to upregulation of phospholipids. • Tombusvirus replication depends on de novo lipid synthesis. • Deletion of FFAT-domain host protein enhances TBSV replication. • TBSV rewires host phospholipid synthesis.« less

Masitinib (AB1010), a Potent and Selective Tyrosine Kinase Inhibitor Targeting KIT

PubMed Central

Dubreuil, Patrice; Letard, Sébastien; Ciufolini, Marco; Gros, Laurent; Humbert, Martine; Castéran, Nathalie; Borge, Laurence; Hajem, Bérengère; Lermet, Anne; Sippl, Wolfgang; Voisset, Edwige; Arock, Michel; Auclair, Christian; Leventhal, Phillip S.; Mansfield, Colin D.; Moussy, Alain; Hermine, Olivier

2009-01-01

Background The stem cell factor receptor, KIT, is a target for the treatment of cancer, mastocytosis, and inflammatory diseases. Here, we characterise the in vitro and in vivo profiles of masitinib (AB1010), a novel phenylaminothiazole-type tyrosine kinase inhibitor that targets KIT. Methodology/Principal Findings In vitro, masitinib had greater activity and selectivity against KIT than imatinib, inhibiting recombinant human wild-type KIT with an half inhibitory concentration (IC50) of 200±40 nM and blocking stem cell factor-induced proliferation and KIT tyrosine phosphorylation with an IC50 of 150±80 nM in Ba/F3 cells expressing human or mouse wild-type KIT. Masitinib also potently inhibited recombinant PDGFR and the intracellular kinase Lyn, and to a lesser extent, fibroblast growth factor receptor 3. In contrast, masitinib demonstrated weak inhibition of ABL and c-Fms and was inactive against a variety of other tyrosine and serine/threonine kinases. This highly selective nature of masitinib suggests that it will exhibit a better safety profile than other tyrosine kinase inhibitors; indeed, masitinib-induced cardiotoxicity or genotoxicity has not been observed in animal studies. Molecular modelling and kinetic analysis suggest a different mode of binding than imatinib, and masitinib more strongly inhibited degranulation, cytokine production, and bone marrow mast cell migration than imatinib. Furthermore, masitinib potently inhibited human and murine KIT with activating mutations in the juxtamembrane domain. In vivo, masitinib blocked tumour growth in mice with subcutaneous grafts of Ba/F3 cells expressing a juxtamembrane KIT mutant. Conclusions Masitinib is a potent and selective tyrosine kinase inhibitor targeting KIT that is active, orally bioavailable in vivo, and has low toxicity. PMID:19789626

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.

Identification of Chemical-Genetic Interactions via Parallel Analysis of Barcoded Yeast Strains.

PubMed

Suresh, Sundari; Schlecht, Ulrich; Xu, Weihong; Miranda, Molly; Davis, Ronald W; Nislow, Corey; Giaever, Guri; St Onge, Robert P

2016-09-01

The Yeast Knockout Collection is a complete set of gene deletion strains for the budding yeast, Saccharomyces cerevisiae In each strain, one of approximately 6000 open-reading frames is replaced with a dominant selectable marker flanked by two DNA barcodes. These barcodes, which are unique to each gene, allow the growth of thousands of strains to be individually measured from a single pooled culture. The collection, and other resources that followed, has ushered in a new era in chemical biology, enabling unbiased and systematic identification of chemical-genetic interactions (CGIs) with remarkable ease. CGIs link bioactive compounds to biological processes, and hence can reveal the mechanism of action of growth-inhibitory compounds in vivo, including those of antifungal, antibiotic, and anticancer drugs. The chemogenomic profiling method described here measures the sensitivity induced in yeast heterozygous and homozygous deletion strains in the presence of a chemical inhibitor of growth (termed haploinsufficiency profiling and homozygous profiling, respectively, or HIPHOP). The protocol is both scalable and amenable to automation. After competitive growth of yeast knockout collection cultures, with and without chemical inhibitors, CGIs can be identified and quantified using either array- or sequencing-based approaches as described here. © 2016 Cold Spring Harbor Laboratory Press.

Induction of a non-specific permeability transition in mitochondria from Yarrowia lipolytica and Dipodascus (Endomyces) magnusii yeasts.

PubMed

Kovaleva, Mariya V; Sukhanova, Evgeniya I; Trendeleva, Tatyana A; Zyl'kova, Marina V; Ural'skaya, Ludmila A; Popova, Kristina M; Saris, Nils-Erik L; Zvyagilskaya, Renata A

2009-06-01

In this study we used tightly-coupled mitochondria from Yarrowia lipolytica and Dipodascus (Endomyces) magnusii yeasts, possessing a respiratory chain with the usual three points of energy conservation. High-amplitude swelling and collapse of the membrane potential were used as parameters for demonstrating induction of the mitochondrial permeability transition due to opening of a pore (mPTP). Mitochondria from Y. lipolytica, lacking a natural mitochondrial Ca(2+) uptake pathway, and from D. magnusii, harboring a high-capacitive, regulated mitochondrial Ca(2+) transport system (Bazhenova et al. J Biol Chem 273:4372-4377, 1998a; Bazhenova et al. Biochim Biophys Acta 1371:96-100, 1998b; Deryabina and Zvyagilskaya Biochemistry (Moscow) 65:1352-1356, 2000; Deryabina et al. J Biol Chem 276:47801-47806, 2001) were very resistant to Ca(2+) overload. However, exposure of yeast mitochondria to 50-100 microM Ca(2+) in the presence of the Ca(2+) ionophore ETH129 induced collapse of the membrane potential, possibly due to activation of the fatty acid-dependent Ca(2+)/nH(+)-antiporter, with no classical mPTP induction. The absence of response in yeast mitochondria was not simply due to structural limitations, since large-amplitude swelling occurred in the presence of alamethicin, a hydrophobic, helical peptide, forming voltage-sensitive ion channels in lipid membranes. Ca(2+)- ETH129-induced activation of the Ca(2+)/H(+)-antiport system was inhibited and prevented by bovine serum albumin, and partially by inorganic phosphate and ATP. We subjected yeast mitochondria to other conditions known to induce the permeability transition in animal mitochondria, i.e., Ca(2+) overload (in the presence of ETH129) combined with palmitic acid (Mironova et al. J Bioenerg Biomembr 33:319-331, 2001; Sultan and Sokolove Arch Biochem Biophys 386:37-51, 2001), SH-reagents, carboxyatractyloside (an inhibitor of the ADP/ATP translocator), depletion of intramitochondrial adenine nucleotide pools

Assessment and characterisation of yeast-based products intended to mitigate ochratoxin exposure using in vitro and in vivo models.

PubMed

Pfohl-Leszkowicz, A; Hadjeba-Medjdoub, K; Ballet, N; Schrickx, J; Fink-Gremmels, J

2015-01-01

The aim of this paper was to evaluate the capacity of several yeast-based products, derived from baker's and brewer's yeasts, to sequester the mycotoxin ochratoxin A (OTA) and to decrease its rate of absorption and DNA adduct formation in vivo. The experimental protocol included in vitro binding studies using isotherm models, in vivo chicken experiments, in which the serum and tissue concentrations of OTA were analysed in the absence and presence of the test compounds, and the profile of OTA-derived metabolites and their associated DNA adducts were determined. Additionally in vitro cell culture studies (HK2 cells) were applied to assess further the effects for yeast cell product enriched with glutathione (GSH) or selenium. Results of the in vitro binding assay in a buffer system indicated the ability of the yeast-based products, as sequester of OTA, albeit at a different level. In the in vitro experiments in chickens, decreased serum and tissue concentrations of treated animals confirmed that yeast-based products are able to prevent the absorption of OTA. A comparison of the binding affinity in a standard in vitro binding assay with the results obtained in an in vivo chicken experiment, however, showed a poor correlation and resulted in a different ranking of the products. More importantly, we could show that yeast-based products actively modulate the biotransformation of OTA in vivo as well as in vitro in a cell culture model. This effect seems to be attributable to residual enzymatic activities in the yeast-based products. An enrichment of yeast cell wall products with GSH or selenium further modulated the profile of the generated OTA metabolites and the associated pattern of OTA-induced DNA adducts by increasing the conversion of OTA into less toxic metabolites such as OTA, OTB and 4-OH-OTA. A reduced absorption and DNA adduct formation was particularly observed with GSH-enriched yeast, whereas selenium-enriched yeasts could counteract the OTA-induced decrease

Effects of Saccharomyces cerevisiae or boulardii yeasts on acute stress induced intestinal dysmotility.

PubMed

West, Christine; Stanisz, Andrew M; Wong, Annette; Kunze, Wolfgang A

2016-12-28

To investigate the capacity of Saccharomyces cerevisiae ( S. cerevisiae ) and Saccharomyces boulardii ( S. boulardii ) yeasts to reverse or to treat acute stress-related intestinal dysmotility. Adult Swiss Webster mice were stressed for 1 h in a wire-mesh restraint to induce symptoms of intestinal dysmotility and were subsequently killed by cervical dislocation. Jejunal and colon tissue were excised and placed within a tissue perfusion bath in which S. cerevisiae , S. boulardii , or their supernatants were administered into the lumen. Video recordings of contractility and gut diameter changes were converted to spatiotemporal maps and the velocity, frequency, and amplitude of propagating contractile clusters (PCC) were measured. Motility pre- and post-treatment was compared between stressed animals and unstressed controls. S. boulardii and S. cerevisiae helped to mediate the effects of stress on the small and large intestine. Restraint stress reduced jejunal transit velocity (mm/s) from 2.635 ± 0.316 to 1.644 ± 0.238, P < 0.001 and jejunal transit frequency (Hz) from 0.032 ± 0.008 to 0.016 ± 0.005, P < 0.001. Restraint stress increased colonic transit velocity (mm/s) from 0.864 ± 0.183 to 1.432 ± 0.329, P < 0.001 and frequency to a lesser degree. Luminal application of S. boulardii helped to restore jejunal and colonic velocity towards the unstressed controls; 1.833 ± 0.688 to 2.627 ± 0.664, P < 0.001 and 1.516 ± 0.263 to 1.036 ± 0.21, P < 0.001, respectively. S. cerevisiae also had therapeutic effects on the stressed gut, but was most apparent in the jejunum. S. cerevisiae increased PCC velocity in the stressed jejunum from 1.763 ± 0.397 to 2.017 ± 0.48, P = 0.0031 and PCC frequency from 0.016 ± 0.009 to 0.027 ± 0.007, P < 0.001. S. cerevisiae decreased colon PCC velocity from 1.647 ± 0.187 to 1.038 ± 0.222, P < 0.001. Addition of S. boulardii or S. cerevisiae supernatants also helped to restore motility to unstressed values in similar capacity

Effects of Saccharomyces cerevisiae or boulardii yeasts on acute stress induced intestinal dysmotility

PubMed Central

West, Christine; Stanisz, Andrew M; Wong, Annette; Kunze, Wolfgang A

2016-01-01

AIM To investigate the capacity of Saccharomyces cerevisiae (S. cerevisiae) and Saccharomyces boulardii (S. boulardii) yeasts to reverse or to treat acute stress-related intestinal dysmotility. METHODS Adult Swiss Webster mice were stressed for 1 h in a wire-mesh restraint to induce symptoms of intestinal dysmotility and were subsequently killed by cervical dislocation. Jejunal and colon tissue were excised and placed within a tissue perfusion bath in which S. cerevisiae, S. boulardii, or their supernatants were administered into the lumen. Video recordings of contractility and gut diameter changes were converted to spatiotemporal maps and the velocity, frequency, and amplitude of propagating contractile clusters (PCC) were measured. Motility pre- and post-treatment was compared between stressed animals and unstressed controls. RESULTS S. boulardii and S. cerevisiae helped to mediate the effects of stress on the small and large intestine. Restraint stress reduced jejunal transit velocity (mm/s) from 2.635 ± 0.316 to 1.644 ± 0.238, P < 0.001 and jejunal transit frequency (Hz) from 0.032 ± 0.008 to 0.016 ± 0.005, P < 0.001. Restraint stress increased colonic transit velocity (mm/s) from 0.864 ± 0.183 to 1.432 ± 0.329, P < 0.001 and frequency to a lesser degree. Luminal application of S. boulardii helped to restore jejunal and colonic velocity towards the unstressed controls; 1.833 ± 0.688 to 2.627 ± 0.664, P < 0.001 and 1.516 ± 0.263 to 1.036 ± 0.21, P < 0.001, respectively. S. cerevisiae also had therapeutic effects on the stressed gut, but was most apparent in the jejunum. S. cerevisiae increased PCC velocity in the stressed jejunum from 1.763 ± 0.397 to 2.017 ± 0.48, P = 0.0031 and PCC frequency from 0.016 ± 0.009 to 0.027 ± 0.007, P < 0.001. S. cerevisiae decreased colon PCC velocity from 1.647 ± 0.187 to 1.038 ± 0.222, P < 0.001. Addition of S. boulardii or S. cerevisiae supernatants also helped to restore motility to unstressed values in similar

Expression profiling reveals an unexpected growth-stimulating effect of surplus iron on the yeast Saccharomyces cerevisiae.

PubMed

Du, Yang; Cheng, Wang; Li, Wei-Fang

2012-08-01

Iron homeostasis plays a crucial role in growth and division of cells in all kingdoms of life. Although yeast iron metabolism has been extensively studied, little is known about the molecular mechanism of response to surplus iron. In this study, expression profiling of Saccharomyces cerevisiae in the presence of surplus iron revealed a dual effect at 1 and 4 h. A cluster of stress-responsive genes was upregulated via activation of the stress-resistance transcription factor Msn4, which indicated the stress effect of surplus iron on yeast metabolism. Genes involved in aerobic metabolism and several anabolic pathways are also upregulated in iron-surplus conditions, which could significantly accelerate yeast growth. This dual effect suggested that surplus iron might participate in a more complex metabolic network, in addition to serving as a stress inducer. These findings contribute to our understanding of the global response of yeast to the fluctuating availability of iron in the environment.

The neglected nano-specific toxicity of ZnO nanoparticles in the yeast Saccharomyces cerevisiae

PubMed Central

Zhang, Weicheng; Bao, Shaopan; Fang, Tao

2016-01-01

Nanoparticles (NPs) with unique physicochemical properties induce nano-specific (excess) toxicity in organisms compared with their bulk counterparts. Evaluation and consideration of nano-specific toxicity are meaningful for the safe design and environmental risk assessment of NPs. However, ZnO NPs have been reported to lack excess toxicity for diverse organisms. In the present study, the nano-specific toxicity of ZnO NPs was evaluated in the yeast Saccharomyces cerevisiae. Nano-specific toxicity of ZnO NPs was not observed in the wild type yeast. However, the ZnO NPs induced very similar nano-specific toxicities in the three mutants with comparable log Te (particle) values (0.64 vs 0.65 vs 0.62), suggesting that the mutants were more sensitive and specific for the NPs’ nano-specific toxicity. The toxic effects in the yeast were slightly attributable to dissolved zinc ions from the ZnO (nano or bulk) particles. Oxidative damage and mechanical damage contributed to the toxic effect of the ZnO particles. The mechanism of mechanical damage is proposed to be an inherent characteristic underlying the nano-specific toxicity in the mutants. The log Te (particle) was a useful parameter for evaluation of NPs nano-specific toxicity, whereas log Te (ion) efficiently determined the NPs toxicity associated with released ions. PMID:27094203

The neglected nano-specific toxicity of ZnO nanoparticles in the yeast Saccharomyces cerevisiae

NASA Astrophysics Data System (ADS)

Zhang, Weicheng; Bao, Shaopan; Fang, Tao

2016-04-01

Nanoparticles (NPs) with unique physicochemical properties induce nano-specific (excess) toxicity in organisms compared with their bulk counterparts. Evaluation and consideration of nano-specific toxicity are meaningful for the safe design and environmental risk assessment of NPs. However, ZnO NPs have been reported to lack excess toxicity for diverse organisms. In the present study, the nano-specific toxicity of ZnO NPs was evaluated in the yeast Saccharomyces cerevisiae. Nano-specific toxicity of ZnO NPs was not observed in the wild type yeast. However, the ZnO NPs induced very similar nano-specific toxicities in the three mutants with comparable log Te (particle) values (0.64 vs 0.65 vs 0.62), suggesting that the mutants were more sensitive and specific for the NPs’ nano-specific toxicity. The toxic effects in the yeast were slightly attributable to dissolved zinc ions from the ZnO (nano or bulk) particles. Oxidative damage and mechanical damage contributed to the toxic effect of the ZnO particles. The mechanism of mechanical damage is proposed to be an inherent characteristic underlying the nano-specific toxicity in the mutants. The log Te (particle) was a useful parameter for evaluation of NPs nano-specific toxicity, whereas log Te (ion) efficiently determined the NPs toxicity associated with released ions.

The neglected nano-specific toxicity of ZnO nanoparticles in the yeast Saccharomyces cerevisiae.

PubMed

Zhang, Weicheng; Bao, Shaopan; Fang, Tao

2016-04-20

Nanoparticles (NPs) with unique physicochemical properties induce nano-specific (excess) toxicity in organisms compared with their bulk counterparts. Evaluation and consideration of nano-specific toxicity are meaningful for the safe design and environmental risk assessment of NPs. However, ZnO NPs have been reported to lack excess toxicity for diverse organisms. In the present study, the nano-specific toxicity of ZnO NPs was evaluated in the yeast Saccharomyces cerevisiae. Nano-specific toxicity of ZnO NPs was not observed in the wild type yeast. However, the ZnO NPs induced very similar nano-specific toxicities in the three mutants with comparable log Te ((particle)) values (0.64 vs 0.65 vs 0.62), suggesting that the mutants were more sensitive and specific for the NPs' nano-specific toxicity. The toxic effects in the yeast were slightly attributable to dissolved zinc ions from the ZnO (nano or bulk) particles. Oxidative damage and mechanical damage contributed to the toxic effect of the ZnO particles. The mechanism of mechanical damage is proposed to be an inherent characteristic underlying the nano-specific toxicity in the mutants. The log Te ((particle)) was a useful parameter for evaluation of NPs nano-specific toxicity, whereas log Te ((ion)) efficiently determined the NPs toxicity associated with released ions.

Yeast-based biosensors: design and applications.

PubMed

Adeniran, Adebola; Sherer, Michael; Tyo, Keith E J

2015-02-01

Yeast-based biosensing (YBB) is an exciting research area, as many studies have demonstrated the use of yeasts to accurately detect specific molecules. Biosensors incorporating various yeasts have been reported to detect an incredibly large range of molecules including but not limited to odorants, metals, intracellular metabolites, carcinogens, lactate, alcohols, and sugars. We review the detection strategies available for different types of analytes, as well as the wide range of output methods that have been incorporated with yeast biosensors. We group biosensors into two categories: those that are dependent upon transcription of a gene to report the detection of a desired molecule and those that are independent of this reporting mechanism. Transcription-dependent biosensors frequently depend on heterologous expression of sensing elements from non-yeast organisms, a strategy that has greatly expanded the range of molecules available for detection by YBBs. Transcription-independent biosensors circumvent the problem of sensing difficult-to-detect analytes by instead relying on yeast metabolism to generate easily detected molecules when the analyte is present. The use of yeast as the sensing element in biosensors has proven to be successful and continues to hold great promise for a variety of applications. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.

A novel bispecific antibody, S-Fab, induces potent cancer cell killing.

PubMed

Li, Li; He, Ping; Zhou, Changhua; Jing, Li; Dong, Bin; Chen, Siqi; Zhang, Ning; Liu, Yawei; Miao, Ji; Wang, Zhong; Li, Qing

2015-01-01

Bispecific antibodies that engage immune cells to kill cancer cells have been actively studied in cancer immunotherapy. In this study, we present a novel bispecific format, S-Fab, fabricated by linking a single-domain anti-carcinoembryonic antigen VHH to a conventional anti-CD3 Fab. In contrast to most bispecific antibodies, the S-Fab bispecific antibody can be efficiently expressed and purified from bacteria. The purified S-Fab is stable in serum and is able to recruit T cells to drive potent cancer cell killing. In xenograft models, the S-Fab antibody suppresses tumor growth in the presence of human immune cells. Our study suggested that the bispecific S-Fab format can be applied to a wide range of immunotherapies.

19-nor vitamin-D analogs: a new class of potent inhibitors of proliferation and inducers of differentiation of human myeloid leukemia cell lines.

PubMed

Asou, H; Koike, M; Elstner, E; Cambell, M; Le, J; Uskokovic, M R; Kamada, N; Koeffler, H P

1998-10-01

We have studied the in vitro biological activities and mechanisms of action of 1,25-dihydroxyvitamin D3 (1,25D3) and nine potent 1,25D3 analogs on proliferation and differentiation of myeloid leukemia cell lines (HL-60, retinoic acid-resistant HL-60 [RA-res HL-60], NB4 and Kasumi-1). The common novel structural motiff for almost all the analogs included removal of C-19 (19-nor); each also had unsaturation of the side chain. All the compounds were potent; for example, the concentration of analogs producing a 50% clonal inhibition (ED50) ranged between 1 x 10(-9) to 4 x 10(-11) mol/L when using the HL-60 cell line. The most active compound [1, 25(OH)2-16,23E-diene-26-trifluoro-19-nor-cholecalciferol (Ro 25-9716)] had an ED50 of 4 x 10(-11) mol/L; in contrast, the 1,25D3 produced an ED50 of 10(-9) mol/L with the HL-60 target cells. Ro 25-9716 (10(-9) mol/L, 3 days) was a strong inducer of myeloid differentiation because it caused 92% of the HL-60 cells to express CD11b and 75% of these cells to reduce nitroblue tetrazolium (NBT). This compound (10(-8) mol/L, 4 days) also caused HL-60 cells to arrest in the G1 phase of the cell cycle (88% cells in G1 v 48% of the untreated control cells). The p27(kip-1), a cyclin-dependent kinase inhibitor which is important in blocking the cell cycle, was induced more quickly and potently by Ro 25-9716 (10(-7) mol/L, 0 to 5 days) than by 1,25D3, suggesting a possible mechanism by which these analogs inhibit proliferation of leukemic growth. The NB4 promyelocytic leukemia cells cultured with the Ro 25-9716 were also inhibited in their clonal proliferation (ED50, 5 x 10(-11) mol/L) and their expression of CD11b was enhanced (80% positive [10(-9) mol/L, 4 days] v 27% untreated NB4 cells). Moreover, the combination of Ro 25-9716 (10(-9) mol/L) and all-trans retinoic acid (ATRA, 10(-7) mol/L) induced 92% of the NB4 cells to reduce NBT, whereas only 26% of the cells became NBT positive after a similar exposure to the combination of 1,25D3

Stochastic simulations of a synthetic bacteria-yeast ecosystem

PubMed Central

2012-01-01

Background The field of synthetic biology has greatly evolved and numerous functions can now be implemented by artificially engineered cells carrying the appropriate genetic information. However, in order for the cells to robustly perform complex or multiple tasks, co-operation between them may be necessary. Therefore, various synthetic biological systems whose functionality requires cell-cell communication are being designed. These systems, microbial consortia, are composed of engineered cells and exhibit a wide range of behaviors. These include yeast cells whose growth is dependent on one another, or bacteria that kill or rescue each other, synchronize, behave as predator-prey ecosystems or invade cancer cells. Results In this paper, we study a synthetic ecosystem comprising of bacteria and yeast that communicate with and benefit from each other using small diffusible molecules. We explore the behavior of this heterogeneous microbial consortium, composed of Saccharomyces cerevisiae and Escherichia coli cells, using stochastic modeling. The stochastic model captures the relevant intra-cellular and inter-cellular interactions taking place in and between the eukaryotic and prokaryotic cells. Integration of well-characterized molecular regulatory elements into these two microbes allows for communication through quorum sensing. A gene controlling growth in yeast is induced by bacteria via chemical signals and vice versa. Interesting dynamics that are common in natural ecosystems, such as obligatory and facultative mutualism, extinction, commensalism and predator-prey like dynamics are observed. We investigate and report on the conditions under which the two species can successfully communicate and rescue each other. Conclusions This study explores the various behaviors exhibited by the cohabitation of engineered yeast and bacterial cells. The way that the model is built allows for studying the dynamics of any system consisting of two species communicating with one

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.

Biotechnological Applications of Dimorphic Yeasts

NASA Astrophysics Data System (ADS)

Doiphode, N.; Joshi, C.; Ghormade, V.; Deshpande, M. V.

The dimorphic yeasts have the equilibrium between spherical growth (budding) and polarized (hyphal or pseudohyphal tip elongation) which can be triggered by change in the environmental conditions. The reversible growth phenomenon has made dimorphic yeasts as an useful model to understand fungal evolution and fungal differentiation, in general. In nature dimorphism is clearly evident in plant and animal fungal pathogens, which survive and most importantly proliferate in the respective hosts. However, number of organisms with no known pathogenic behaviour also show such a transition, which can be exploited for the technological applications due to their different biochemical make up under different morphologies. For instance, chitin and chitosan production using dimorphic Saccharomyces, Mucor, Rhizopus and Benjaminiella, oil degradation and biotransformation with yeast-form of Yarrowia species, bioremediation of organic pollutants, exopolysac-charide production by yeast-phase of Aureobasidium pullulans, to name a few. Myrothecium verrucaria can be used for seed dressing in its yeast form and it produces a mycolytic enzyme complex in its hyphal-form for the biocontrol of fungal pathogens, while Beauveria bassiana and other entomopathogens kill the insect pest by producing yeast- like cells in the insect body. The form-specific expression of protease, chitinase, lipase, ornithine decarboxylase, glutamate dehydrogenases, etc. make Benjaminiella poitrasii, Basidiobolus sp., and Mucor rouxii strains important in bioremediation, nanobiotechnology, fungal evolution and other areas.

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

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.

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

Combined Stimulation with Interleukin-18 and Interleukin-12 Potently Induces Interleukin-8 Production by Natural Killer Cells.

PubMed

Poznanski, Sophie M; Lee, Amanda J; Nham, Tina; Lusty, Evan; Larché, Margaret J; Lee, Dean A; Ashkar, Ali A

2017-01-01

The combination of interleukin (IL)-18 and IL-12 (IL-18+IL-12) potently stimulates natural killer (NK) cells, triggering an innate immune response to infections and cancers. Strategies exploiting the effects of IL-18+IL-12 have shown promise for cancer immunotherapy. However, studies have primarily characterized the NK cell response to IL-18+IL-12 in terms of interferon (IFN)-γ production, with little focus on other cytokines produced. IL-8 plays a critical role in activating and recruiting immune cells, but it also has tumor-promoting functions. IL-8 is classically produced by regulatory NK cells; however, cytotoxic NK cells do not typically produce IL-8. In this study, we uncover that stimulation with IL-18+IL-12 induces high levels of IL-8 production by ex vivo expanded and freshly isolated NK cells and NK cells in peripheral blood mononuclear cells. We further report that tumor necrosis factor (TNF)-α, produced by NK cells following IL-18+IL-12 stimulation, regulates IL-8 production. The IL-8 produced is in turn required for maximal IFN-γ and TNF-α production. These findings may have important implications for the immune response to infections and cancer immunotherapies. This study broadens our understanding of NK cell function and IL-18+IL-12 synergy by uncovering an unprecedented ability of IL-18+IL-12-activated peripheral blood NK cells to produce elevated levels of IL-8 and identifying the requirement for intermediates induced by IL-18+IL-12 for maximal cytokine production following stimulation. © 2017 S. Karger AG, Basel.

The preclinical biology of a new potent and selective progestin: trimegestone.

PubMed

Winneker, Richard C; Bitran, Daniel; Zhang, Zhiming

2003-11-01

Trimegestone (TMG) is a 19-norpregnane progestin being developed, in combination with an estrogen, for the treatment of postmenopausal symptoms. TMG binds to the human progesterone receptor with an affinity greater than medroxyprogesterone acetate (MPA), norethindrone (NET), and levonorgestrel (LNG). In contrast, TMG binds with low affinity to the androgen, glucocorticoid and mineralocorticoid receptor and has no measurable affinity for the estrogen receptor. Compared to other progestins, TMG demonstrates an improved separation of its PR affinity from its affinity to other classical steroid hormone receptors. In vivo, TMG has potent progestin activity. For example, TMG produces glandular differentiation of the uterine endometrium in rabbits and is about 30 and 60 times more potent than MPA and NET, respectively. In the rat, TMG maintains pregnancy, induces deciduoma formation, inhibits ovulation and has uterine anti-estrogenic activity. With respect to these endpoints, TMG appears to be more potent and selective on uterine epithelial responses than other classical progestin responses. In vivo, TMG does not have significant androgenic, glucocorticoid, anti-glucocorticoid or mineralocorticoid activity but does have anti-mineralocorticoid activity and modest anti-androgenic effects. This overall profile is qualitatively similar to progesterone. When TMG is administered chronically, it antagonizes the effect of estradiol on the uterus but does not antagonize the beneficial bone sparing activity of estradiol. In rat studies evaluating CNS GABAA receptor modulatory activity, TMG is less active on this likely undesirable endpoint than progesterone and norethindrone acetate, which may translate into fewer mood-related side effects. The results indicate that TMG is a potent and selective progestin with a preclinical profile well suited for hormone replacement therapy.

Effect of a new potent CCK antagonist, lorglumide, on caerulein- and bombesin-induced pancreatic secretion and growth in the rat.

PubMed Central

Scarpignato, C.; Varga, G.; Dobronyi, I.; Papp, M.

1989-01-01

1. The effect of lorglumide, a new potent cholecystokinin (CCK) antagonist, on pancreatic secretion and growth induced by caerulein and bombesin was studied in the rat. 2. Pancreatic exocrine secretion was studied both in vitro (isolated and perfused pancreatic segments) and in vivo (anaesthetized animals with cannulation of the common bile duct) whereas the trophic effect was investigated after short-term (5 days) administration of the peptides and/or lorglumide. 3. Both caerulein and bombesin stimulated amylase release from in vitro pancreatic segments in a concentration-dependent manner. Although the efficacy of both peptides was virtually identical, the potency of caerulein was higher than that of bombesin. Lorglumide displaced the concentration-response curves to caerulein to the right without affecting the maximum response, suggesting a competitive antagonism. The Schild plot analysis of data gave a straight line with a slope not significantly different from unity. The calculated pA2 for lorglumide was 7.31 +/- 0.45. The antagonist, however, was completely ineffective when tested against bombesin-induced amylase release. 4. In vivo experiments confirmed results from in vitro studies since lorglumide (5 and 10 mg kg-1) significantly reduced pancreatic exocrine secretion induced by caerulein without affecting the response to bombesin. 5. Administration of either peptide increased the weight of the pancreas, the total pancreatic protein and DNA, trypsin and amylase content. Lorglumide (10 mg kg-1), administered together with caerulein, reduced the peptide-induced increase in pancreatic weight, protein and enzyme content. On the contrary, when lorglumide was given together with bombesin, all the parameters that were examined were not altered by concomitant administration of the antagonist.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:2470456

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

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

Evaluation of Automated Yeast Identification System

NASA Technical Reports Server (NTRS)

McGinnis, M. R.

1996-01-01

One hundred and nine teleomorphic and anamorphic yeast isolates representing approximately 30 taxa were used to evaluate the accuracy of the Biolog yeast identification system. Isolates derived from nomenclatural types, environmental, and clinica isolates of known identity were tested in the Biolog system. Of the isolates tested, 81 were in the Biolog database. The system correctly identified 40, incorrectly identified 29, and was unable to identify 12. Of the 28 isolates not in the database, 18 were given names, whereas 10 were not. The Biolog yeast identification system is inadequate for the identification of yeasts originating from the environment during space program activities.

Potent Immune Responses in Rhesus Macaques Induced by Nonviral Delivery of a Self-amplifying RNA Vaccine Expressing HIV Type 1 Envelope With a Cationic Nanoemulsion

PubMed Central

Bogers, Willy M.; Oostermeijer, Herman; Mooij, Petra; Koopman, Gerrit; Verschoor, Ernst J.; Davis, David; Ulmer, Jeffrey B.; Brito, Luis A.; Cu, Yen; Banerjee, Kaustuv; Otten, Gillis R.; Burke, Brian; Dey, Antu; Heeney, Jonathan L.; Shen, Xiaoying; Tomaras, Georgia D.; Labranche, Celia; Montefiori, David C.; Liao, Hua-Xin; Haynes, Barton; Geall, Andrew J.; Barnett, Susan W.

2015-01-01

Self-amplifying messenger RNA (mRNA) of positive-strand RNA viruses are effective vectors for in situ expression of vaccine antigens and have potential as a new vaccine technology platform well suited for global health applications. The SAM vaccine platform is based on a synthetic, self-amplifying mRNA delivered by a nonviral delivery system. The safety and immunogenicity of an HIV SAM vaccine encoding a clade C envelope glycoprotein formulated with a cationic nanoemulsion (CNE) delivery system was evaluated in rhesus macaques. The HIV SAM vaccine induced potent cellular immune responses that were greater in magnitude than those induced by self-amplifying mRNA packaged in a viral replicon particle (VRP) or by a recombinant HIV envelope protein formulated with MF59 adjuvant, anti-envelope binding (including anti-V1V2), and neutralizing antibody responses that exceeded those induced by the VRP vaccine. These studies provide the first evidence in nonhuman primates that HIV vaccination with a relatively low dose (50 µg) of formulated self-amplifying mRNA is safe and immunogenic. PMID:25234719

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

Behavior of yeast cells in aqueous suspension affected by pulsed electric field.

PubMed

El Zakhem, H; Lanoisellé, J-L; Lebovka, N I; Nonus, M; Vorobiev, E

2006-08-15

This work discusses pulsed electric fields (PEF) induced effects in treatment of aqueous suspensions of concentrated yeast cells (S. cerevisiae). The PEF treatment was done using pulses of near-rectangular shape, electric field strength was within E=2-5 kV/cm and the total time of treatment was t(PEF)=10(-4)-0.1 s. The concentration of aqueous yeast suspensions was in the interval of C(Y)=0-22 (wt%), where 1% concentration corresponds to the cellular density of 2x10(8) cells/mL. Triton X-100 was used for studying non-ionic surfactant additive effects. The electric current peak value I was measured during each pulse application, and from these data the electrical conductivity sigma was estimated. The PEF-induced damage results in increase of sigma with t(PEF) increasing and attains its saturation level sigma approximately sigma(max) at long time of PEF treatment. The value of sigma(max) reflects the efficiency of damage. The reduced efficiency of damage at suspension volume concentration higher than phi(Y) approximately 32 vol% is explained by the percolation phenomenon in the randomly packed suspension of near-spherical cells. The higher cytoplasmic ions leakage was observed in presence of surfactant. Experiments were carried out in the static and continuous flow treatment chambers in order to reveal the effects of mixing in PEF-treatment efficiency. A noticeable aggregation of the yeast cells was observed in the static flow chamber during the PEF treatment, while aggregation was not so pronounced in the continuous flow chamber. The nature of the enhanced aggregation under the PEF treatment was revealed by the zeta-potential measurements: these data demonstrate different zeta-potential signs for alive and dead cells. The effect of the electric field strength on the PEF-induced extraction of the intracellular components of S. cerevisiae is discussed.

The discovery of thienopyridine analogues as potent IkappaB kinase beta inhibitors. Part II.

PubMed

Wu, Jiang-Ping; Fleck, Roman; Brickwood, Janice; Capolino, Alison; Catron, Katrina; Chen, Zhidong; Cywin, Charles; Emeigh, Jonathan; Foerst, Melissa; Ginn, John; Hrapchak, Matt; Hickey, Eugene; Hao, Ming-Hong; Kashem, Mohammed; Li, Jun; Liu, Weimin; Morwick, Tina; Nelson, Richard; Marshall, Daniel; Martin, Leslie; Nemoto, Peter; Potocki, Ian; Liuzzi, Michel; Peet, Gregory W; Scouten, Erika; Stefany, David; Turner, Michael; Weldon, Steve; Zimmitti, Clare; Spero, Denise; Kelly, Terence A

2009-10-01

An SAR study that identified a series of thienopyridine-based potent IkappaB Kinase beta (IKKbeta) inhibitors is described. With focuses on the structural optimization at C4 and C6 of structure 1 (Fig. 1), the study reveals that small alkyl and certain aromatic groups are preferred at C4, whereas polar groups with proper orientation at C6 efficiently enhance compound potency. The most potent analogues inhibit IKKbeta with IC50s as low as 40 nM, suppress LPS-induced TNF-alpha production in vitro and in vivo, display good kinase selectivity profiles, and are active in a HeLa cell NF-kappaB reporter gene assay, demonstrating that they directly interfere with the NF-kappaB signaling pathway.

A pilot study on seborrheic dermatitis using pramiconazole as a potent oral anti-Malassezia agent.

PubMed

Piérard, Gérald E; Ausma, Jannie; Henry, Frédérique; Vroome, Valérie; Wouters, Luc; Borgers, Marcel; Cauwenbergh, Geert; Piérard-Franchimont, Claudine

2007-01-01

Seborrheic dermatitis is considered to be a Malassezia-driven disease. Little objective information is available so far from biometrological quantitative assessments of this skin condition. Pramiconazole is a novel triazole with potent in vitro antifungal activity, especially against Malassezia spp. To study the sequential effects of pramiconazole on Malassezia, inflammation and epidermal changes. This study was performed in 2 groups of subjects suffering from seborrheic dermatitis. The first group (n = 17) remained untreated and was used as control. Clinical, mycological and biometrological assessments were performed at inclusion and during the following 2 weeks. The second group of subjects (n = 10) received a single 200-mg oral dose of pramiconazole at inclusion. Clinical, mycological and biometrological evaluations were performed before and during 1 month following the single antifungal intake. For both parts of the study, several parameters were assessed including yeast density, desquamation, erythema, itching and sebum excretion. In the control group, no significant changes were observed in any of the parameters during the observation period. The findings were markedly different in the pramiconazole-treated subjects. The yeast density was significantly improved on days 3, 7 and 28. Desquamation, erythema, itching, and the global clinical evaluation as assessed by the patients and investigators became significantly improved on days 7 and 28. A trend in decrease of scaliness was noted. No effect on sebum excretion was evidenced. In conclusion, a single 200-mg dose of pramiconazole exhibitsin vivo efficacy in controlling some important clinical aspects of seborrheic dermatitis. Following a reduction in the number of yeasts on day 3, a decrease in the severity of clinical signs and symptoms occurred from day 7 onwards. Sebum excretion appeared uninvolved in the clearing process of seborrheic dermatitis. A single 200-mg dose of pramiconazole appears to abate

Genomics and the making of yeast biodiversity.

PubMed

Hittinger, Chris Todd; Rokas, Antonis; Bai, Feng-Yan; Boekhout, Teun; Gonçalves, Paula; Jeffries, Thomas W; Kominek, Jacek; Lachance, Marc-André; Libkind, Diego; Rosa, Carlos A; Sampaio, José Paulo; Kurtzman, Cletus P

2015-12-01

Yeasts are unicellular fungi that do not form fruiting bodies. Although the yeast lifestyle has evolved multiple times, most known species belong to the subphylum Saccharomycotina (syn. Hemiascomycota, hereafter yeasts). This diverse group includes the premier eukaryotic model system, Saccharomyces cerevisiae; the common human commensal and opportunistic pathogen, Candida albicans; and over 1000 other known species (with more continuing to be discovered). Yeasts are found in every biome and continent and are more genetically diverse than angiosperms or chordates. Ease of culture, simple life cycles, and small genomes (∼10-20Mbp) have made yeasts exceptional models for molecular genetics, biotechnology, and evolutionary genomics. Here we discuss recent developments in understanding the genomic underpinnings of the making of yeast biodiversity, comparing and contrasting natural and human-associated evolutionary processes. Only a tiny fraction of yeast biodiversity and metabolic capabilities has been tapped by industry and science. Expanding the taxonomic breadth of deep genomic investigations will further illuminate how genome function evolves to encode their diverse metabolisms and ecologies. Copyright © 2015 Elsevier Ltd. All rights reserved.

Allyl m-Trifluoromethyldiazirine Mephobarbital: An Unusually Potent Enantioselective and Photoreactive Barbiturate General Anesthetic

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

Savechenkov, Pavel Y.; Zhang, Xi; Chiara, David C.

2012-12-10

We synthesized 5-allyl-1-methyl-5-(m-trifluoromethyl-diazirynylphenyl)barbituric acid (14), a trifluoromethyldiazirine-containing derivative of general anesthetic mephobarbital, separated the racemic mixture into enantiomers by chiral chromatography, and determined the configuration of the (+)-enantiomer as S by X-ray crystallography. Additionally, we obtained the {sup 3}H-labeled ligand with high specific radioactivity. R-(-)-14 is an order of magnitude more potent than the most potent clinically used barbiturate, thiopental, and its general anesthetic EC{sub 50} approaches those for propofol and etomidate, whereas S-(+)-14 is 10-fold less potent. Furthermore, at concentrations close to its anesthetic potency, R-(-)-14 both potentiated GABA-induced currents and increased the affinity for the agonist muscimol inmore » human {alpha}1{beta}2/3{gamma}2L GABA{sub A} receptors. Finally, R-(-)-14 was found to be an exceptionally efficient photolabeling reagent, incorporating into both {alpha}1 and {beta}3 subunits of human {alpha}1{beta}3 GABAA receptors. These results indicate R-(-)-14 is a functional general anesthetic that is well-suited for identifying barbiturate binding sites on Cys-loop receptors.« less

The sweet taste of death: glucose triggers apoptosis during yeast chronological aging.

PubMed

Ruckenstuhl, Christoph; Carmona-Gutierrez, Didac; Madeo, Frank

2010-10-01

As time goes by, a postmitotic cell ages following a degeneration process ultimately ending in cell death. This phenomenon is evolutionary conserved and present in unicellular eukaryotes as well, making the yeast chronological aging system an appreciated model. Here, single cells die in a programmed fashion (both by apoptosis and necrosis) for the benefit of the whole population. Besides its meaning for aging and cell death research, age-induced programmed cell death represents the first experimental proof for the so-called group selection theory: Apoptotic genes became selected during evolution because of the benefits they might render to the whole cell culture and not to the individual cell. Many anti‐aging stimuli have been discovered in the yeast chronological aging system and have afterwards been confirmed in higher cells or organisms. New work from the Burhans group (this issue) now demonstrates that glucose signaling has a progeriatric effect on chronologically aged yeast cells: Glucose administration results in a diminished efficacy of cells to enter quiescence, finally causing superoxide‐mediated replication stress and apoptosis.

[Genetic system for maintaining the mitochondrial human genome in yeast Yarrowia lipolytica].

PubMed

Isakova, E P; Deryabina, Yu I; Velyakova, A V; Biryukova, J K; Teplova, V V; Shevelev, A B

2016-01-01

For the first time, the possibility of maintaining an intact human mitochondrial genome in a heterologous system in the mitochondria of yeast Yarrowia lipolytica is shown. A method for introducing directional changes into the structure of the mitochondrial human genome replicating in Y. lipolytica by an artificially induced ability of yeast mitochondria for homologous recombination is proposed. A method of introducing and using phenotypic selection markers for the presence or absence of defects in genes tRNA-Lys and tRNA-Leu of the mitochondrial genome is developed. The proposed system can be used to correct harmful mutations of the human mitochondrial genome associated with mitochondrial diseases and for preparative amplification of intact mitochondrial DNA with an adjusted sequence in yeast cells. The applicability of the new system for the correction of mutations in the genes of Lys- and Leu-specific tRNAs of the human mitochondrial genome associated with serious and widespread human mitochondrial diseases such as myoclonic epilepsy with lactic acidosis (MELAS) and myoclonic epilepsy with ragged-red fibers (MERRF) is shown.

Strong and multi-antigen specific immunity by hepatitis B core antigen (HBcAg)-based vaccines in a murine model of chronic hepatitis B: HBcAg is a candidate for a therapeutic vaccine against hepatitis B virus.

PubMed

Akbar, Sheikh Mohammad Fazle; Chen, Shiyi; Al-Mahtab, Mamun; Abe, Masanori; Hiasa, Yoichi; Onji, Morikazu

2012-10-01

Experimental evidence suggests that hepatitis B core antigen (HBcAg)-specific cytotoxic T lymphocytes (CTL) are essential for the control of hepatitis B virus (HBV) replication and prevention of liver damage in patients with chronic hepatitis B (CHB). However, most immune therapeutic approaches in CHB patients have been accomplished with hepatitis B surface antigen (HBsAg)-based prophylactic vaccines with unsatisfactory clinical outcomes. In this study, we prepared HBsAg-pulsed dendritic cells (DC) and HBcAg-pulsed DC by culturing spleen DC from HBV transgenic mice (HBV TM) and evaluated the immunomodulatory capabilities of these antigens, which may serve as a better therapy for CHB. The kinetics of HBsAg, antibody levels against HBsAg (anti-HBs), proliferation of HBsAg- and HBcAg-specific lymphocytes, production of antigen-specific CTL, and activation of endogenous DC were compared between HBV TM vaccinated with either HBsAg- or HBcAg-pulsed DC. Vaccination with HBsAg-pulsed DC induced HBsAg-specific immunity, but failed to induce HBcAg-specific immunity in HBV TM. However, immunization of HBV TM with HBcAg-pulsed DC resulted in: (1) HBsAg negativity, (2) production of anti-HBs, and (3) development of HBsAg- and HBcAg-specific T cells and CTL in the spleen and the liver. Additionally, significantly higher levels of activated endogenous DC were detected in HBV TM immunized with HBcAg-pulsed DC compared to HBsAg-pulsed DC (p<0.05). The capacity of HBcAg to modulate both HBsAg- and HBcAg-specific immunity in HBV TM, and activation of endogenous DC in HBV TM without inducing liver damage suggests that HBcAg should be an integral component of the therapeutic vaccine against CHB. Copyright © 2012 Elsevier B.V. All rights reserved.

[Groups and sources of yeasts in house dust].

PubMed

Glushakova, A M; Zheltikova, T M; Chernov, I Iu

2004-01-01

House dust contains bacteria, mycelial fungi, microarthropods, and yeasts. The house dust samples collected in 25 apartments in Moscow and the Moscow region were found to contain yeasts belonging to the genera Candida, Cryptococcus, Debaryomyces, Rhodotorula, Sporobolomyces, and Trichosporon. The most frequently encountered microorganisms were typical epiphytic yeasts, such as Cryptococcus diffluens and Rhodotorula mucilaginosa, which are capable of long-term preservation in an inactive state. The direct source of epiphytic yeasts occurring in the house dust might be the indoor plants, which were contaminated with these yeasts, albeit to a lesser degree than outdoor plants. Along with the typical epiphytic yeasts, the house dust contained the opportunistic yeast pathogens Candida catenulata, C. guillermondii, C. haemulonii, C. rugosa, and C. tropicalis, which are known as the causal agents of candidiasis. We failed to reveal any correlation between the abundance of particular yeast species in the house dust, residential characteristics, and the atopic dermatitis of the inhabitants.

Genomics and the making of yeast biodiversity

USDA-ARS?s Scientific Manuscript database

Yeasts are unicellular fungi that do not form fruiting bodies. Although the yeast lifestyle has evolved multiple times, most known species belong to the subphylum Saccharomycotina (syn. Hemiascomycota, hereafter yeasts). This diverse group includes the premier eukaryotic model system, Saccharomyces ...

Development of a new benzophenone-diketopiperazine-type potent antimicrotubule agent possessing a 2-pyridine structure.

PubMed

Hayashi, Yoshiki; Takeno, Haruka; Chinen, Takumi; Muguruma, Kyohei; Okuyama, Kohei; Taguchi, Akihiro; Takayama, Kentaro; Yakushiji, Fumika; Miura, Masahiko; Usui, Takeo; Hayashi, Yoshio

2014-10-09

A new benzophenone-diketopiperazine-type potent antimicrotubule agent was developed by modifying the structure of the clinical candidate plinabulin (1). Although the right-hand imidazole ring with a branched alkyl chain at the 5-position in 1 was critical for the potency of the antimicrotubule activity, we successfully substituted this moiety with a simpler 2-pyridyl structure by converting the left-hand ring from a phenyl to a benzophenone structure without decreasing the potency. The resultant compound 6b (KPU-300) exhibited a potent cytotoxicity, with an IC50 value of 7.0 nM against HT-29 cells, by strongly binding to tubulin (K d = 1.3 μM) and inducing microtubule depolymerization.

Aromatic Amino Acid-Derived Compounds Induce Morphological Changes and Modulate the Cell Growth of Wine Yeast Species

PubMed Central

González, Beatriz; Vázquez, Jennifer; Cullen, Paul J.; Mas, Albert; Beltran, Gemma; Torija, María-Jesús

2018-01-01

Yeasts secrete a large diversity of compounds during alcoholic fermentation, which affect growth rates and developmental processes, like filamentous growth. Several compounds are produced during aromatic amino acid metabolism, including aromatic alcohols, serotonin, melatonin, and tryptamine. We evaluated the effects of these compounds on growth parameters in 16 different wine yeasts, including non-Saccharomyces wine strains, for which the effects of these compounds have not been well-defined. Serotonin, tryptamine, and tryptophol negatively influenced yeast growth, whereas phenylethanol and tyrosol specifically affected non-Saccharomyces strains. The effects of the aromatic alcohols were observed at concentrations commonly found in wines, suggesting a possible role in microbial interaction during wine fermentation. Additionally, we demonstrated that aromatic alcohols and ethanol are able to affect invasive and pseudohyphal growth in a manner dependent on nutrient availability. Some of these compounds showed strain-specific effects. These findings add to the understanding of the fermentation process and illustrate the diversity of metabolic communication that may occur among related species during metabolic processes. PMID:29696002

21 CFR 172.590 - Yeast-malt sprout extract.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Yeast-malt sprout extract. 172.590 Section 172.590... Substances § 172.590 Yeast-malt sprout extract. Yeast-malt sprout extract, as described in this section, may... produced by partial hydrolysis of yeast extract (derived from Saccharomyces cereviseae, Saccharomyces...

Yeasts Diversity in Fermented Foods and Beverages

NASA Astrophysics Data System (ADS)

Tamang, Jyoti Prakash; Fleet, Graham H.

People across the world have learnt to culture and use the essential microorganisms for production of fermented foods and alcoholic beverages. A fermented food is produced either spontaneously or by adding mixed/pure starter culture(s). Yeasts are among the essential functional microorganisms encountered in many fermented foods, and are commercially used in production of baker's yeast, breads, wine, beer, cheese, etc. In Asia, moulds are predominant followed by amylolytic and alcohol-producing yeasts in the fermentation processes, whereas in Africa, Europe, Australia and America, fermented products are prepared exclusively using bacteria or bacteria-yeasts mixed cultures. This chapter would focus on the varieties of fermented foods and alcoholic beverages produced by yeasts, their microbiology and role in food fermentation, widely used commercial starters (pilot production, molecular aspects), production technology of some common commercial fermented foods and alcoholic beverages, toxicity and food safety using yeasts cultures and socio-economy

Organotins Are Potent Activators of PPARγ and Adipocyte Differentiation in Bone Marrow Multipotent Mesenchymal Stromal Cells

PubMed Central

Yanik, Susan C.; Baker, Amelia H.; Mann, Koren K.; Schlezinger, Jennifer J.

2011-01-01

Adipocyte differentiation in bone marrow is potentially deleterious to both bone integrity and lymphopoiesis. Here, we examine the hypothesis that organotins, common environmental contaminants that are dual ligands for peroxisome proliferator–activated receptor (PPAR) γ and its heterodimerization partner retinoid X receptor (RXR), are potent activators of bone marrow adipogenesis. A C57Bl/6-derived bone marrow multipotent mesenchymal stromal cell (MSC) line, BMS2, was treated with rosiglitazone, a PPARγ agonist, bexarotene, an RXR agonist, or a series of organotins. Rosiglitazone and bexarotene potently activated adipocyte differentiation; however, bexarotene had a maximal efficacy of only 20% of that induced by rosiglitazone. Organotins (tributyltin [TBT], triphenyltin, and dibutyltin) also stimulated adipocyte differentiation (EC50 of 10–20nM) but with submaximal, structure-dependent efficacy. In coexposures, both bexarotene and TBT enhanced rosiglitazone-induced adipogenesis. To investigate the contribution of PPARγ to TBT-induced adipogenesis, we examined expression of PPARγ2, as well as its transcriptional target FABP4. TBT-induced PPARγ2 and FABP4 protein expression with an efficacy intermediate between rosiglitazone and bexarotene, similar to lipid accumulation. A PPARγ antagonist and PPARγ-specific small hairpin RNA suppressed TBT-induced differentiation, although to a lesser extent than rosiglitazone-induced differentiation, suggesting that TBT may engage alternate pathways. TBT and bexarotene, but not rosiglitazone, also induced the expression of TGM2 (an RXR target) and ABCA1 (a liver X receptor target). The results show that an environmental contaminant, acting with the same potency as a therapeutic drug, induces PPARγ-dependent adipocyte differentiation in bone marrow MSCs. Activation of multiple nuclear receptor pathways by organotins may have significant implications for bone physiology. PMID:21622945

Targeting the genital tract mucosa with a lipopeptide/recombinant adenovirus prime/boost vaccine induces potent and long-lasting CD8+ T cell immunity against herpes: importance of MyD88.

PubMed

Zhang, Xiuli; Dervillez, Xavier; Chentoufi, Aziz Alami; Badakhshan, Tina; Bettahi, Ilham; Benmohamed, Lbachir

2012-11-01

Targeting of the mucosal immune system of the genital tract with subunit vaccines has failed to induce potent and durable local CD8(+) T cell immunity, which is crucial for protection against many sexually transmitted viral pathogens, including HSV type 2 (HSV-2), which causes genital herpes. In this study, we aimed to investigate the potential of a novel lipopeptide/adenovirus type 5 (Lipo/rAdv5) prime/boost mucosal vaccine for induction of CD8(+) T cell immunity to protect the female genital tract from herpes. The lipopeptide vaccine and the rAdv5 vaccine express the immunodominant HSV-2 CD8(+) T cell epitope (gB(498-505)), and both were delivered intravaginally in the progesterone-induced B6 mouse model of genital herpes. Compared with mice immunized with the homologous lipopeptide/lipopeptide (Lipo/Lipo) vaccine, the Lipo/rAdv5 prime/boost immunized mice 1) developed potent and sustained HSV-specific CD8(+) T cells, detected in both the genital tract draining nodes and in the vaginal mucosa; 2) had significantly lower virus titers; 3) had decreased overt signs of genital herpes disease; and 4) did not succumb to lethal infection (p < 0.005) after intravaginal HSV-2 challenge. Polyfunctional CD8(+) T cells, producing IFN-γ, TNF-α, and IL-2 and exhibiting cytotoxic activity, were associated with protection (p < 0.005). The protective CD8(+) T cell response was significantly compromised in the absence of the adapter MyD88 (p = 0.0001). Taken together, these findings indicate that targeting of the vaginal mucosa with a Lipo/rAdv5 prime/boost vaccine elicits a potent, MyD88-dependent, and long-lasting mucosal CD8(+) T cell protective immunity against sexually transmitted herpes infection and disease.

Genetics of Yeasts

NASA Astrophysics Data System (ADS)

Querol, Amparo; Fernández-Espinar, M. Teresa; Belloch, Carmela

The use of yeasts in biotechnology processes dates back to ancient days. Before 7000 BC, beer was produced in Sumeria. Wine was made in Assyria in 3500 BC, and ancient Rome had over 250 bakeries, which were making leavened bread by 100 BC. And milk has been made into Kefyr and Koumiss in Asia for many centuries (Demain, Phaff, & Kurtzman, 1999). However, the importance of yeast in the food and beverage industries was only realized about 1860, when their role in food manufacturing became evident.

Application of computerised penile arterial waveform analysis in the diagnosis of arteriogenic impotence. An initial study in potent and impotent men.

PubMed

Desai, K M; Gingell, J C; Skidmore, R; Follett, D H

1987-11-01

A new method is described for evaluating arteriogenic impotence by means of noninvasive quantification of penile Doppler arterial waveforms using computerised analysis based on the Laplace Transform model. The haemodynamic changes occurring during a papaverine-induced erection in healthy potent volunteers have been recorded by this technique, which has also been shown to be capable of discriminating between a normal and an abnormal penile arterial supply in an initial study of potent and impotent men.

21 CFR 172.590 - Yeast-malt sprout extract.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Yeast-malt sprout extract. 172.590 Section 172.590... CONSUMPTION Flavoring Agents and Related Substances § 172.590 Yeast-malt sprout extract. Yeast-malt sprout... prescribed conditions: (a) The additive is produced by partial hydrolysis of yeast extract (derived from...

21 CFR 172.590 - Yeast-malt sprout extract.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Yeast-malt sprout extract. 172.590 Section 172.590... CONSUMPTION Flavoring Agents and Related Substances § 172.590 Yeast-malt sprout extract. Yeast-malt sprout... prescribed conditions: (a) The additive is produced by partial hydrolysis of yeast extract (derived from...

21 CFR 172.590 - Yeast-malt sprout extract.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Yeast-malt sprout extract. 172.590 Section 172.590... CONSUMPTION Flavoring Agents and Related Substances § 172.590 Yeast-malt sprout extract. Yeast-malt sprout... prescribed conditions: (a) The additive is produced by partial hydrolysis of yeast extract (derived from...