Gene–gene and gene–environment interactions in complex traits in yeast

One of the fundamental questions in biology is how the genotype regulates the phenotype. An increasing number of studies indicate that, in most cases, the effect of a genetic locus on the phenotype is context‐dependent, i.e. it is influenced by the genetic background and the environment in which the phenotype is measured. Still, the majority of the studies, in both model organisms and humans, that map the genetic regulation of phenotypic variation in complex traits primarily identify additive loci with independent effects. This does not reflect an absence of the contribution of genetic interactions to phenotypic variation, but instead is a consequence of the technical limitations in mapping gene–gene interactions (GGI) and gene–environment interactions (GEI). Yeast, with its detailed molecular understanding, diverse population genomics and ease of genetic manipulation, is a unique and powerful resource to study the contributions of GGI and GEI in the regulation of phenotypic variation. Here we review studies in yeast that have identified GGI and GEI that regulate phenotypic variation, and discuss the contribution of these findings in explaining missing heritability of complex traits, and how observations from these GGI and GEI studies enhance our understanding of the mechanisms underlying genetic robustness and adaptability that shape the architecture of the genotype–phenotype map.     

The frenemies within: viruses,retrotransposons and plasmids that naturally infect Saccharomyces yeasts

Viruses are a major focus of current research efforts because of their detrimental impact on humanity and their ubiquity within the environment. Bacteriophages have long been used to study host–virus interactions within microbes, but it is often forgotten that the single‐celled eukaryote Saccharomyces cerevisiae and related species are infected with double‐stranded RNA viruses, single‐stranded RNA viruses, LTR‐retrotransposons and double‐stranded DNA plasmids. These intracellular nucleic acid elements have some similarities to higher eukaryotic viruses, i.e. yeast retrotransposons have an analogous lifecycle to retroviruses, the particle structure of yeast totiviruses resembles the capsid of reoviruses and segregation of yeast plasmids is analogous to segregation strategies used by viral episomes. The powerful experimental tools available to study the genetics, cell biology and evolution of S. cerevisiae are well suited to further our understanding of how cellular processes are hijacked by eukaryotic viruses, retrotransposons and plasmids. This article has been written to briefly introduce viruses, retrotransposons and plasmids that infect Saccharomyces yeasts, emphasize some important cellular proteins and machineries with which they interact, and suggest the evolutionary consequences of these interactions. Copyright © 2017 John Wiley & Sons, Ltd.     

Paracoccin from Paracoccidioides brasiliensis; purification through affinity with chitin and identification of N‐acetyl‐β‐D‐glucosaminidase activity

The dimorphic fungus Paracoccidioides brasiliensis is the causative agent of paracoccidioidomycosis, the most frequent systemic mycosis in Latin America. Our group has been working with paracoccin, a P. brasiliensis lectin with MM 70 kDa, which is purified by affinity with immobilized N‐acetylglucosamine (GlcNAc). Paracoccin has been described to play a role in fungal adhesion to extracellular matrix components and to induce high and persistent levels of TNFα and nitric oxide production by macrophages. In the cell wall, paracoccin colocalizes with the β‐1,4‐homopolymer of GlcNAc into the budding sites of the P. brasiliensis yeast cell. In this paper we present a protocol for the chitin‐affinity purification of paracoccin. This procedure provided higher yields than those achieved by means of the technique based on the affinity of this lectin with GlcNAc and had an impact on downstream assays. SDS–PAGE and Western blot analysis revealed similarities between the N‐acetylglucosamine‐ and chitin‐bound fractions, confirmed by MALDI–TOF–MS of trypsinic peptides. Western blot of two‐dimensional gel electrophoresis of the yeast extract showed a major spot with M_r 70 000 and pI approximately 5.63. Morevover, an N‐acetyl‐β‐D ‐glucosaminidase activity was reported for paracoccin, thereby providing new insights into the mechanisms that lead to cell wall remodelling and opening new perspectives for its structural characterization. Copyright © 2009 John Wiley & Sons, Ltd.     

Curcumin provides potential protection against the activation of hypoxia and prolyl 4-hydroxylase inhibitors on prostate-specific antigen expression in human prostate carcinoma cells

Scope: Prostate‐specific antigen (PSA) is a well‐known marker for diagnosing and monitoring prostate cancer. Curcumin, a yellow curry pigment, has been reported to enhance androgen receptor (AR) degradation. We examined the effects of curcumin on increasing PSA expression by hypoxia and prolyl hydroxylase inhibitors, L ‐mimosine and dimethyloxalylglycine (DMOG), in human prostate carcinoma LNCaP cells. Methods and results: The ³H‐thymidine incorporation assay revealed that either L ‐mimosine or DMOG treatments attenuated cell proliferation. Immunoblot and enzyme‐linked immunosorbent assays (ELISA) indicated that both L ‐mimosine and DMOG have an effect similar to hypoxia, which stabilized hypoxia‐inducible factor‐1α (HIF‐1α) and induced PSA gene expression. The results of the immunoblot and transient gene expression assays indicated that induction of the PSA expression by hypoxia is both HIF‐1α‐ and AR‐dependent. Immunoblot assays revealed that a curcumin treatment (10 μM) decreased the protein abundance of AR but did not significantly affect the protein levels of HIF‐1α and vascular endothelial growth factor, which were induced by hypoxia. ELISA and transient gene expression assays indicated that curcumin blocked the activation of L ‐mimosine or DMOG treatment on PSA expression. Conclusions: These results indicate that curcumin blocked the enhanced effect of PSA expression by L ‐mimosine and DMOG that induce hypoxia condition.     

Starch–lipid and starch–lipid–protein complexes: A comprehensive review

Physical interactions often occur between major food components during food processing. These interactions may involve starch, lipids, and proteins forming V‐type starch–lipid complexes or ternary starch–lipid–protein complexes of larger molecular size and greater structural order. Complexes between starch and lipids have been the subject of intensive research for over half a century, whereas the study of starch–lipid–protein complexes is a relatively new field with only a limited amount of knowledge being gained so far. The formation of these complexes can significantly affect the functional and nutritional properties of finished food products in terms of flavor, texture, shelf life, and digestibility. This article provides a comprehensive review of starch–lipid and starch–lipid–protein complexes, including their classification, factors affecting their formation and structure, and preparative and analytical methods. The review also considers how complexes affect the physicochemical and functional properties of starch, including digestibility, and potential applications in the food industry.     

Characterization of α‐tubulin gene distinctively presented in a cytoplasmic male sterile and its maintainer line of non‐heading Chinese cabbage

BACKGROUND: Microtubules are prominent components of the cytoskeleton in every eukaryotic cell. Plant microtubules are essential for a wide variety of cellular functions, including generation of cell polarity, intracellular transport, positioning of organelles, cell wall deposition and cell division. The major component of microtubules is tubulin, an α,β heterodimer protein with a molecular mass of each subunit of around 50 kDa. Tubulin exists in cells as a mixture of polypeptides differing in their isoelectric points. Some post‐translational modifications of tubulins are thought to modulate the functions and localization of microtubules within the cell. RESULTS: The complete sequence of a single‐copy α‐tubulin gene Tuba1, belonging to a multiple gene family of non‐heading Chinese cabbage (Brassica campestris ssp. chinensis Makino), was obtained. The gene was expressed in high levels in young leaves and stamens, and it was also highly expressed during all stages of microsporogenesis in the maintainer. However, there was a distinct difference in α‐tubulin expression between the sterile stage and the normal stages of pollen in a cytoplasmic male sterility line and its maintainer. CONCLUSION: Tuba1 was significantly related to the cell division and elongation of non‐heading Chinese cabbage, demonstrating that this gene played an important role in the development of pollen and may be closely related to male sterility. Copyright © 2008 Society of Chemical Industry     

Constitutive Effects of Lead on Aryl Hydrocarbon Receptor Gene Battery and Protection by β‐carotene and Ascorbic Acid in Human HepG2 Cells

Lead (Pb) is an environmental pollutant that can get entry into human body through contaminated foods, drinks, and inhaled air leading to severe biological consequences, and has been responsible for many deaths worldwide. The objectives of this study were 1st to investigate the modulatory effects of environmentally relevant concentrations of Pb on AhR gene battery, which is controlling xenobiotics metabolism. 2nd, trials to reduce Pb‐induced adverse effects were done using some phytochemicals like β‐carotene or ascorbic acid. Human hepatoma (HepG2) cell lines were exposed to a wide range of Pb concentrations varying from physiological to toxic levels (0 to 10 mg/L) for 24 h. High Pb concentrations (1 to 10 mg/L) significantly reduced phase I (CYP1A1 and 1A2) and phase II (UGT1A6 and NQO1) xenobiotic metabolizing enzyme mRNA expression in a mechanistic manner through the AhR regulation pathway. Additionally, these Pb concentrations induced oxidative stress in HepG2 cells in terms of production of reactive oxygen species (ROS) and induced heme oxygenase‐1 mRNA expression in a concentration‐dependent phenomenon. Coexposure of HepG2 cells to physiological concentrations of some micronutrients, like β‐carotene (10 μM) or ascorbic acid (0.1 mM), along with Pb (1 mg/L) for 24 h significantly reduced the levels of ROS production and recovered AhR mRNA expression into the normal levels. Thus, consumption of foods rich in these micronutrients may help to reduce the adverse effects of lead in areas with high levels of pollution.     

Selection of reference genes for quantitative real‐time PCR in postharvest tomatoes (Lycopersicon esculentum) treated by continuous low‐voltage direct current electricity to increase secondary metabolites

Quantitative real‐time PCR (RT‐qPCR) is often used for gene expression analysis to reveal molecular mechanism of how stresses can enhance the secondary metabolites production. For RT‐qPCR to be valid, robust reference genes are required. This study validated nine candidate genes as reference genes using BestKeeper, NormFinder and geNorm methods in RT‐qPCR analysis of postharvest tomatoes subjected to electricity‐induced stress. The most stable genes as indicated by each method were EF‐1α by BestKeeper; CAC by NormFinder; and PP2Acs/TIP41 by geNorm. Due to the inconsistency in the ranking of the candidate genes by the three methods, the pairwise variation from geNorm analysis was used to calculate the minimum numbers of reference genes for an accurate normalisation and revealed that a combination of PP2Acs and TIP41 was an optimum. This reference gene combination was further validated for their stability in RT‐qPCR analysis of four carotenoid‐related genes, and more reliable expression levels were obtained.     

Detection of Proteinases in Saccharomyces cerevisiae by Flow Cytometry

The physiological state of a yeast population used for inoculation determines how rapidly the cells adapt to new environmental conditions, begin proliferating and utilising extract. The decision as to whether a yeast culture is suitable for re‐pitching should not be based only on viability determinations since this can be misleading. Increased proteolytic activity in a yeast population indicates the onset of senescence. A flow cytometric method has been developed for measuring a wide variety of proteinases in Saccharomyces cerevisiae employing a commercially available casein‐dye conjugate. The detection of intracellular proteinase activity gives an early indication of apoptotic events and allows improved assessment of the physiological state of a yeast population. This knowledge will assist the industry to optimize the selection of yeast and its subsequent fermentation performance. Yeast cell autolysis with all its negative consequences for beer quality and stability will thus be minimised.     

Targeting carbon for crop yield and drought resilience

Current methods of crop improvement are not keeping pace with projected increases in population growth. Breeding, focused around key traits of stem height and disease resistance, delivered the step‐change yield improvements of the green revolution of the 1960s. However, subsequently, yield increases through conventional breeding have been below the projected requirement of 2.4% per year required by 2050. Genetic modification (GM) mainly for herbicide tolerance and insect resistance has been transformational, akin to a second green revolution, although GM has yet to make major inroads into intrinsic yield processes themselves. Drought imposes the major restriction on crop yields globally but, as yet, has not benefited substantially from genetic improvement and still presents a major challenge to agriculture. Much still has to be learnt about the complex process of how drought limits yield and what should be targeted. Mechanisms of drought adaptation from the natural environment cannot be taken into crops without significant modification for the agricultural environment because mechanisms of drought tolerance are often in contrast with mechanisms of high productivity required in agriculture. However, through convergence of fundamental and translational science, it would appear that a mechanism of sucrose allocation in crops can be modified for both productivity and resilience to drought and other stresses. Recent publications show how this mechanism can be targeted by GM, natural variation and a new chemical approach. Here, with an emphasis on drought, we highlight how understanding fundamental science about how crops grow, develop and what limits their growth and yield can be combined with targeted genetic selection and pioneering chemical intervention technology for transformational yield improvements. © 2017 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

Lipid metabolic effect of Korean red ginseng extract in mice fed on a high-fat diet

BACKGROUND: Ginseng saponin and ginsenosides exert anti‐obesity effects via the modulation of physiological lipid metabolism in vivo or intracellular signalling in cell culture systems. However, the complicated relationship between the anti‐obesity effects of ginseng and gene expression has yet to be defined under in vivo conditions. Therefore, we evaluated the relationship between the anti‐obesity effects of Korean red ginseng extract (KRGE) and hepatic gene expression profiles in mice fed long‐term on a high‐fat diet (HFD) in this study. RESULTS: KRGE reduces the levels of cholesterol, low‐density lipoprotein‐cholesterol (LDL‐C), serum triglycerides, and atherogenic indices. Levels of leptin, adiponectin and insulin, which regulate glucose and lipid metabolism, were impaired profoundly by HFD. However, KRGE treatment brought these levels back to normal. KRGE was found to down‐regulate genes associated with lipid metabolism or cholesterol metabolism (Lipa, Cyp7a1, Il1rn, Acot2, Mogat1, Osbpl3, Asah3l, Insig1, Anxa2, Vldlr, Hmgcs1, Sytl4, Plscr4, Pla2g4e, Slc27a3, Enpp6), all of which were up‐regulated by HFD. CONCLUSION: KRGE regulated the expression of genes associated with abnormal physiology via HFD. Leptin, insulin, and adiponectin, which carry out critical functions in energy and lipid metabolism, were shown to be modulated by KRGE. These results show that KRGE is effective in preventing obesity. Copyright © 2011 Society of Chemical Industry     

Genome‐wide expression profiling of the osmoadaptation response of Debaryomyces hansenii

The euryhaline marine yeast Debaromyces hansenii is a model system for the study of processes related to osmoadaptation. In this study, microarray‐based gene expression analyses of the entire genome of D. hansenii was used to study its response to osmotic stress. Differential gene expression, compared to control, was examined at three time points (0.5, 3 and 6 h) after exposure of D. hansenii cultures to high salt concentration. Among the 1.72% of genes showing statistically significant differences in expression, only 65 genes displayed at least three‐fold increases in mRNA levels after treatment with 2 M NaCl. On the other hand, 44 genes showed three‐fold repression. Upregulated as well as the downregulated genes were grouped into functional categories to identify biochemical processes possibly affected by osmotic stress and involved in osmoadaptation. The observation that only a limited number of genes are upregulated in D. hansenii in response to osmotic stress supports the notion that D. hansenii is pre‐adapted to survive in extreme saline environments. In addition, since more than one‐half of the upregulated genes encode for ribosomal proteins, it is possible that a translational gene regulatory mechanism plays a key role in D. hansenii's osmoregulatory response. Validation studies for ENA1 and for hyphal wall/cell elongation protein genes, using real‐time PCR, confirmed patterns of gene expression observed in our microarray experiments. To our knowledge, this study is the first of its kind in this organism and provides the foundation for future molecular studies assessing the significance of the genes identified here in D. hansenii's osmoadaptation. Copyright © 2009 John Wiley & Sons, Ltd.     

Effective suppression of azoxymethane‐induced aberrant crypt foci formation in mice with citrus peel flavonoids

Citrus peel or its extract has been reported to exhibit a broad spectrum of biological activity. Herein, we report the first investigation of inhibitory effects of a formulated product from citrus peel extract, gold lotion (GL), on azoxymethane‐induced colonic tumorigenesis. We have demonstrated that oral feeding of GL decreased the number of aberrant crypt foci (ACF), particularly large size of ACF in colonic tissues of mice. Both gene and protein expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase‐2 (COX‐2) were suppressed by GL treatment. The in vivo data have revealed for the first time that the citrus peel extract–GL–is an effective antitumor agent mechanistically downregulating the protein levels of iNOS, COX‐2, ornithine decarboxylase, vascular endothelial growth factor, and matrix metallopeptidase 9 in colonic tissues of mice, suggesting that GL is a novel functional natural product capable of preventing inflammation‐associated colon tumorigenesis.     

Application of pure and mixed probiotic lactic acid bacteria and yeast cultures for oat fermentation

Fermentation of a prebiotic containing oat substrate with probiotic lactic acid bacteria and yeast strains is an intriguing approach for the development of new synbiotic functional products. This approach was applied in the present work by using pure and mixed microbial cultures to ferment a heat‐treated oat mash. Results show that the strains studied were appropriate for oat fermentation and the process could be completed for 6–10 h depending on the strain. The viable cell counts achieved within this time were above the required levels of 10⁶–10⁷ cfu ml⁻¹ for probiotic products. Both single lactic acid bacteria strains and mixed cultures of the same strains with yeast were found suitable for oat fermentation. However, the pure LAB cultures attributed better flavour and shelf life of the oat drinks. The content of the prebiotic oat component beta‐glucan remained within 0.30–0.36% during fermentation and storage of the drinks obtained with each of the strains used. Thus, these products would contribute diet with the valuable functional properties of beta‐glucan. Also, the viability of pure and mixed cultures in the oat products was good: levels of cell counts remained above the required numbers for probiotic products throughout the estimated shelf‐life period. Copyright © 2005 Society of Chemical Industry