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No abstract is available for this article. [ABSTRACT FROM AUTHOR]

During the last couple of decades, the methylotrophic yeast, Pichia pastoris, has emerged as an important yeast species owing to its increasing importance both in industry and in basic research. The presently available methods for P. pastoris transformation necessitate the preparation of competent cells, which requires lots of resource, space, time, and efforts. This limits the number of transformations that can be performed by an individual in a given time. This paper is reporting a modification in the available protocols, which makes P. pastoris transformation hassle‐free. In the present, modified procedure, cells were grown in patches on YPD plate(s), and the rest of the steps were carried out in small Eppendorf tubes. This modified protocol does not require a big centrifuge and shaker. This modified procedure of P. pastoris transformation with its unique way of competent cells preparation will be helpful for those working with this yeast species. [ABSTRACT FROM AUTHOR]

All authors of the present paper have worked in labs that participated to the sequencing effort of the Saccharomyces cerevisiae reference genome, and we owe to this the fact that we have all chosen to work on genomics of yeasts. S. cerevisiae has been a popular model species for genetics since the 20th century as well as being a model for general eukaryotic cellular processes. Although it has also been used empirically in fermentation for millennia, there was until recently, a lack of knowledge about the natural and evolutionary history of this yeast. The achievement of the international effort to sequence its genome was the foundation for understanding many eukaryotic biological processes but also represented the first step towards the study of the genome and ecological diversity of yeast populations worldwide. We will describe recent advances in yeast comparative and population genomics that find their origins in the S. cerevisiae genome project initiated and pursued by André Goffeau. [ABSTRACT FROM AUTHOR]

No abstract is available for this article. [ABSTRACT FROM AUTHOR]

No abstract is available for this article. [ABSTRACT FROM AUTHOR]

Abstract: In the last decade several collections of Saccharomyces cerevisiae yeast strains have been created. In these collections every gene is modified in a similar manner such as by a deletion or the addition of a protein tag. Such libraries have enabled a diversity of systematic screens, giving rise to large amounts of information regarding gene functions. However, often papers describing such screens focus on a single gene or a small set of genes and all other loci affecting the phenotype of choice (‘hits’) are only mentioned in tables that are provided as supplementary material and are often hard to retrieve or search. To help unify and make such data accessible, we have created a Database of High Throughput Screening Hits (dHITS). The dHITS database enables information to be obtained about screens in which genes of interest were found as well as the other genes that came up in that screen – all in a readily accessible and downloadable format. The ability to query large lists of genes at the same time provides a platform to easily analyse hits obtained from transcriptional analyses or other screens. We hope that this platform will serve as a tool to facilitate investigation of protein functions to the yeast community. [ABSTRACT FROM AUTHOR]

The last two years have seen the publication of three genome-wide gene expression studies of the fission yeast cell cycle. While these microarray papers largely agree on the main patterns of cell cycle-regulated transcription and its control, there are discrepancies with regard to the identity and numbers of periodically expressed genes. We present benchmark and reproducibility analyses showing that the main discrepancies do not reflect differences in the data themselves (microarray or synchronization methods seem to lead only to minor biases) but rather in the interpretation of the data. Our reanalysis of the three datasets reveals that combining all independent information leads to an improved identification of periodically expressed genes. These evaluations suggest that the available microarray data do not allow reliable identification of more than about 500 cell cycle-regulated genes. The temporal expression pattern of the top 500 periodically expressed genes is generally consistent across experiments and the three studies, together with our integrated analysis, provide a coherent and rich source of information on cell cycle-regulated gene expression in Schizosaccharomyces pombe. The reanalysed datasets and other supplementary information are available from an accompanying website: . We hope that this paper will resolve the apparent discrepancies between the previous studies and be useful both for wet-lab biologists and for theoretical scientists who wish to take advantage of the data for follow-up work. Copyright © 2006 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Fission yeast is a powerful model organism that has provided insights into important cellular processes thanks to the ease of its genome editing by homologous recombination. However, creation of strains with a large number of targeted mutations or containing plasmids has been challenging because only a very small number of selection markers is available in Schizosaccharomyces pombe. In this paper, we identify two fission yeast fluoride exporter channels (Fex1p and Fex2p) and describe the development of a new strategy using Fex1p as a selection marker for transformants in rich media supplemented with fluoride. To our knowledge this is the first positive selection marker identified in S. pombe that does not use auxotrophy or drug resistance and that can be used for plasmids transformation or genomic integration in rich media. We illustrate the application of our new marker by significantly accelerating the protocol for genome edition using CRISPR/Cas9 in S. pombe. Copyright © 2016 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The present paper describes the behaviour of Lachancea thermotolerans and Saccharomyces cerevisiae in pure, co-cultured and sequential fermentations in cv. Emir grape must. Faster fermentation rates were observed in wine made with a pure culture of S. cerevisiae and wine produced with simultaneously inoculated cultures of L. thermotolerans and S. cerevisiae. Both L. thermotolerans and S. cerevisiae gave high population numbers. The use of L. thermotolerans in mixed and sequential cultures led to an increase in final total acidity content in the wines, varying in the range 5.40-6.28 g/l (as tartaric acid), compared to pure culture S. cerevisiae, which gave the lowest level of total acidity (5 g/l). The increase was in the order of 1.18-2.06 g/l total acidity. Increase in final acidity by the use of L. thermotolerans might be useful to improve wines with low acidity due to global climate change. Volatile acidity levels (as acetic acid) were in the range 0.53-0.73 g/l, while the concentration of ethyl alcohol varied in the range 10.76-11.62% v/v. Sequential fermentations of wines and pure culture fermentation of L. thermotolerans resulted in reduction in the concentrations of acetaldehyde and higher alcohols, with exception of N-propanol and esters. According to the sensory analysis, wine obtained with sequential inoculation of L. thermotolerans followed by inoculation of S. cerevisiae after 24 h, and simultaneous inoculation of these yeasts, was the most preferred. Copyright © 2016 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Green fluorescent protein (GFP) has become an invaluable tool in biological research. Many GFP variants have been created that differ in brightness, photostability, and folding robustness. We have created two hybrid GFP variants, Envy and Ivy, which we placed in a vector for the C-terminal tagging of yeast proteins by PCR-mediated recombination. The Envy GFP variant combines mutations found in the robustly folding SuperfolderGFP and GFP γ, while the Ivy GFP variant is a hybrid of GFP γ and the yellow-green GFP variant, Clover. We compared Envy and Ivy to EGFP, SuperfolderGFP and GFP γ and found that Envy is brighter than the other GFP variants at both 30°C and 37°C, while Ivy is the most photostable. Envy and Ivy are recognized by a commonly used anti-GFP antibody, and both variants can be immunoprecipitated using the GFP TRAP Camelidae antibody nanotrap technology. Because Envy is brighter than the other GFP variants and is as photostable as GFP γ, we suggest that Envy should be the preferred GFP variant, while Ivy may be used in cases where photostability is of the utmost importance. The GenBank Accession No. for Envy is KM891731, for Ivy is KM891732 and for the yeast optimized GFP γ described in this paper is KM891733. Copyright © 2015 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

In this paper, we report for the first time that Candida melibiosica 2491 yeast strain expresses enhanced phytase activity when used as a biocatalyst in biofuel cells. The polarization also results in an increase of the yeast biomass. Higher steady-state electrical outputs, assigned to earlier production of an endogenous mediator, were achieved at continuous polarization under constant load. The obtained results prove that the C. melibiosica yeast-based biofuel cell could be used for simultaneous electricity generation and phytate bioremediation. In addition, the higher phytase activity obtained by interruptive polarization suggests a new method for increasing the phytase yield from microorganisms. Copyright © 2014 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

This paper describes the results obtained by analysing the human pathogen Candida albicans cell wall subproteome by mass spectrometry, using extraction procedures aimed at releasing proteins bound by disulphide bridges (RAE-CWP) or alkali-labile ester linkages (ALS-CWP). Ten of the total proteins released from the wall by β-ME and/or NaOH contained a potential signal peptide, lacked a GPI cell wall hydrophobic C-terminal domain and were identified as true wall proteins by in silico analysis, whereas four additional proteins were identified as bound to the plasma membrane. The results surprisingly demonstrated that, in addition to the expected RAE-CWP and ALS-CWP proteins, 16 GPI proteins were bound to the wall by disulphide or alkali-sensitive bonds, since they were released by β-ME and/or NaOH. The biological significance of these results is discussed in relation to the added complexity of the organization of the proteins in the C. albicans cell wall. Copyright © 2014 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

This paper developed a novel strategy to improve the fluorescence in situ hybridization-flow cytometry (FISH-FCM) enumeration performance in filamentous yeast species in activated sludge by snailase partial digestion to fully disaggregate filamentous yeast chains into single cells. A 2 h 2% snailase partial digestion liberated more rod-shaped yeast single cells from intertwined filamentous yeast samples than did sonication disaggregation, based on an optical microscopic observation and the forward-light-scatter frequency histogram of FCM analysis. However, adding snailase resulted in a fluorescence-quenching phenomenon of the hybridized filamentous yeast cells, which was minimized by lowering the snailase concentration. An approximately 3 h 0.5% snailase partial digestion conducted between sonication and hybridization significantly improved the FISH-FCM enumeration performance for filamentous yeast species by 37%. The results presented here will facilitate the rapid detection, identification and exact enumeration of specific filamentous fungal species in environmental samples. Copyright © 2012 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

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 Mr 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. [ABSTRACT FROM AUTHOR]

In a previous paper we reported the construction of a S. cerevisiae strain lacking the essential gene LSM4, which could survive by the introduction of a truncated form of the orthologous gene from Kluyveromyces lactis. This strain showed apoptotic hallmarks and other phenotypes, including an increased sensitivity to caffeine and acetic acid. The suppression of the latter phenotype by overexpressing yeast genes allowed the isolation of PGK1, the gene encoding the glycolytic enzyme phosphoglycerate kinase. This gene restored normal ageing, oxygen peroxide resistance and nuclear integrity in the mutant. Other phenotypes, such as caffeine sensitivity and glycerol utilization, were also suppressed. Copyright © 2009 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The coordination between cell wall synthesis and cell growth in the yeast Saccharomyces cerevisiae implicates the PKC1-dependent MAP kinase pathway. KNR4, encoding a 505 amino acid long protein, participates in this coordination, since it displays synthetic lethality with all the members of the PKC1 pathway and shows physical interaction with Slt2/Mpk1. The recent finding that KNR4 interacts genetically or physically with more than 100 partners implicated in different cellular processes raised the question of how these interactions may occur and their physiological significance. This called for an in-depth structure-function analysis of the Knr4 protein, which is reported in the present paper. Computational analysis supported by biochemical and biophysical data characterize Knr4 as a newly identified member of the growing family of intrinsically disordered proteins. Despite disordered regions that are located at the N- and C-termini and are probably responsible for fine regulatory function; this protein contains a structured central core (amino acid residues 80-340) that is able to restore wild-type phenotypes of knr4Δ mutant in stress conditions. However, this fragment was unable to complement synthetic lethality between knr4 mutations and deletions of genes encoding protein kinases of the PKC1-dependent pathway. For these crucial events to occur, the presence of the N-terminal part of Knr4 protein is indispensable. Moreover, we demonstrate that this protein is essential for cell viability in the absence of a functional Pkc1-Slt2 pathway, since the lethality caused by KNR4 deletion in such a genetic background could not be compensated by overexpression of any gene from yeast genomic libraries. Copyright © 2008 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The dimorphic fungus Paracoccidioides brasiliensis is the causative agent of the most frequent systemic mycosis in Latin America. In humans, infection starts by inhalation of fungal propagules, which reach the pulmonary epithelium and differentiate into the yeast parasitic phase. Here we describe the characterization of a Dfg5p ( defective for filamentous growth) homologue of P. brasiliensis, a predictable cell wall protein, first identified in Saccharomyces cerevisiae. The protein, the cDNA and genomic sequences were analysed. The cloned cDNA was expressed in Escherichia coli and the purified r PbDfg5p was used to obtain polyclonal antibodies. Immunoelectron microscopy and biochemical studies demonstrated the presence of PbDfg5p in the fungal cell wall. Enzymatic treatments identified PbDfg5p as a β-glucan linked protein that undergoes N-glycosylation. The r PbDfg5p bound to extracellular matrix components, indicating that those interactions could be important for initial steps leading to P. brasiliensis attachment and colonization of host tissues. The P. brasiliensis dfg5 nucleotide and deduced protein, PbDfg5p, sequences reported in this paper had been submitted to the GenBank database under Accession Nos AY307855 (cDNA) and DQ534495 (genomic). Copyright © 2007 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

It has been proposed that a bottom-fermenting yeast strain of Saccharomyces pastorianus is a natural hybrid between S. cerevisiae and S. bayanus and possesses at least two types of genome. In the process of conducting expressed sequence tag (EST) analysis, we isolated bottom- fermenting yeast-specific ( BFY) genes that have no significant homology with sequences in the S288C database. One of the BFY genes, AMI1, encodes a protein with homology to an amidase conserved among plants, Bacillus subtilis, Neurospora crassa, Schizosaccharomyces pombe and Saccharomyces species, with the exception of S. cerevisiae S288C. In the bottom-fermenting yeast, three alleles of AMI1 (one AMI1-A and two AMI1-B alleles) were found on different chromosomes. AMI1-A on chromosome XIII is most homologous to the S. bayanus AMI1 gene, while AMI1-B on chromosome X is most homologous to the Saccharomyces paradoxus AMI1 gene. Overproduction of AMI1 in S. cerevisiae resulted in a slow-growth phenotype. Although a hydropathy plot shows that Ami1p has a putative signal sequence, it was located in the cell, not secreted into the medium. By metabolome analysis of intracellular compounds, the amount of histidine and arginine is increased, and the amount of threonine, lysine and nicotinic acid is decreased in the Ami1p-overproducing strain as compared with the control, suggesting that Ami1p may hydrolyse some amides related to amino acid and niacin metabolism in the cell. The nucleotide sequence data reported in this paper appear in the DDBJ/EMBL/GenBank nucleotide database under Accession Nos AB195821 and AB195822. Copyright © 2007 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The yeast Saccharomyces cerevisiae is an important industrial microorganism. Nowadays, it is being used as a cell factory for the production of pharmaceuticals such as insulin, although this yeast has long been utilized in the bakery to raise dough, and in the production of alcoholic beverages, fermenting the sugars derived from rice, wheat, barley, corn and grape juice. S. cerevisiae has also been extensively used as a model eukaryotic system. In the last decade, genomic techniques have revealed important features of its molecular biology. For example, DNA array technologies are routinely used for determining gene expression levels in cells under different physiological conditions or environmental stimuli. Laboratory strains of S. cerevisiae are different from wine strains. For instance, laboratory yeasts are unable to completely transform all the sugar in the grape must into ethanol under winemaking conditions. In fact, standard culture conditions are usually very different from winemaking conditions, where multiple stresses occur simultaneously and sequentially throughout the fermentation. The response of wine yeasts to these stimuli differs in some aspects from laboratory strains, as suggested by the increasing number of studies in functional genomics being conducted on wine strains. In this paper we review the most recent applications of post-genomic techniques to understand yeast physiology in the wine industry. We also report recent advances in wine yeast strain improvement and propose a reference framework for integration of genomic information, bioinformatic tools and molecular biology techniques for cellular and metabolic engineering. Finally, we discuss the current state and future perspectives for using 'modern' biotechnology in the wine industry. Copyright © 2007 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The enzyme glyoxylate reductase reversibly reduces glyoxylate to glycolate, or alternatively hydroxypyruvate to D-glycerate, using either NADPH or NADH as a co-factor. The enzyme has multiple metabolic roles in different organisms. In this paper we show that GOR1 (ORF YNL274c) encodes a glyoxylate reductase and not a hydroxyisocaproate dehydrogenase in Saccharomyces cerevisiae, even though it also has minor activity on α-ketoisocaproate. In addition, we show that deletion of the glyoxylate reductase-encoding gene leads to higher biomass concentration after diauxic shift. Copyright © 2006 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

In all eukaryotes, the alignment of the mitotic spindle with the axis of cell polarity is essential for accurate chromosome segregation as well as for the establishment of cell fate, and thus morphogenesis, during development. Studies in invertebrates, higher eukaryotes and yeast suggest that astral microtubules interact with the cell cortex to position the spindle. These microtubules are thought to impose pushing or pulling forces on the spindle poles to affect the rotation or movement of the spindle. In the fission yeast model, where cell division is symmetrical, spindle rotation is dependent on the interaction of astral microtubules with the cortical actin cytoskeleton. In these cells, a bub1-dependent mitotic checkpoint, the spindle orientation checkpoint (SOC), is activated when the spindles fail to align with the cell polarity axis. In this paper we review the mechanism that orientates the spindle during mitosis in fission yeast, and discuss the consequences of misorientation on metaphase progression. Copyright © 2006 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

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In this paper we describe molecular characterization of the TIM9 gene encoding the essential mitochondrial inner-membrane protein in the methylotrophic yeast Pichia methanolica. PmTIM9 contains two exons corresponding to a gene product of 89 amino acid residues and a 140 bp intron. The deduced amino acid sequence exhibited high identity to those of other yeast Tim9ps, and possessed two CX3C motifs that contained two cysteine residues conserved among small Tim family proteins. Moreover, PmTIM9 had the ability to partially suppress the temperature sensitivity of Saccharomyces cerevisiae strain tim9-3, suggesting that PmTIM9 is a functional homologue of the ScTIM9 gene. Copyright © 2006 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Systems biology focuses on obtaining a quantitative description of complete biological systems, even complete cellular function. In this way, it will be possible to perform computer-guided design of novel drugs, advanced therapies for treatment of complex diseases, and to perform in silico design of advanced cell factories for production of fuels, chemicals, food ingredients and pharmaceuticals. The yeast Saccharomyces cerevisiae represents an excellent model system; the density of biological information available on this organism allows it to serve as a eukaryotic model for studying human diseases. Furthermore, it serves as an industrial workhorse for production of a wide range of chemicals and pharmaceuticals. Systems biology involves the combination of novel experimental techniques from different disciplines as well as functional genomics, bioinformatics and mathematical modelling, and hence no single laboratory has access to all the necessary competences. For this reason the Yeast Systems Biology Network (YSBN) has been established. YSBN will coordinate research efforts in yeast systems biology and, through the recently obtained EU funding for a Coordination Action, it will be possible to set appropriate guidelines, establish an appropriate infrastructure for the network and organize courses, meetings and conferences that will consolidate the network and promote systems biology. This paper discusses the impacts of systems biology and how YSBN may play a role in the future development of the field. Copyright © 2006 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Purine salvage is a complex pathway allowing a correct balance between adenylic and guanylic derivatives. In this paper, we show that GUD1 ( YDL238c) encodes guanine deaminase, a catabolic enzyme producing xanthine and ammonia from guanine. Importantly, Gud1p activity was higher during post-diauxic growth, suggesting that a decrease of the guanylic nucleotide pool could be required when cells shift from proliferation to quiescence. Copyright © 2004 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

In this paper, we describe molecular characterization of the FLD1 gene, which encodes glutathione-dependent formaldehyde dehydrogenase (FLD), from the methylotrophic yeast Pichia methanolica. The P. methanolica FLD1 gene contains two exons corresponding to a gene product of 380 amino acid residues and a 225 bp intron, respectively, and its deduced amino acid sequence shows high similarity to those of Fld1ps from other methylotrophic yeasts (80-88%). In P. methanolica, FLD activity is mainly induced by methanol, and this induction is not completely repressed by glucose. Moreover, the expression of the PmFLD1 is strictly regulated, mainly at the mRNA level, its expression increasing with increasing methanol concentrations in the medium. These results suggest that FLD1 is involved in the detoxification of formaldehyde in methanol metabolism, and Fld1p coordinates the formaldehyde level in methanol-grown cells according to the methanol concentration on growth. The sequence has been submitted to the DDBJ/EMBL/GenBank databases under Accession No. AB112935. Copyright © 2004 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Saccharomyces cerevisiae is a facultative anaerobe devoid of mitochondrial alternative oxidase. In this yeast, the structure and biogenesis of the respiratory chain, on the one hand, and the functional interactions of oxidative phosphorylation with the cellular energetic metabolism, on the other, are well documented. However, to our knowledge, the molecular aspects and the physiological roles of the non-respiratory pathways that utilize molecular oxygen have not yet been reviewed. In this paper, we review the various non-respiratory pathways in a global context of utilization of molecular oxygen in S. cerevisiae. The roles of these pathways are examined as a function of environmental conditions, using either physiological, biochemical or molecular data. Special attention is paid to the characterization of the so-called ‘cyanide-resistant respiration’ that is induced by respiratory deficiency, catabolic repression and oxygen limitation during growth. Finally, several aspects of oxygen sensing are discussed. Copyright © 2003 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

In this paper, we present evidence that there are two closely linked copies of the ATP3 gene coding for the γ subunit of the F1F0-ATPase complex (EC3.6.1.34) in four laboratory strains of Saccharomyces cerevisiae, even though the yeast genome project has reported that ATP3 is a single-copy gene on chromosome II. We previously reported that the gene dosage (three copies) of ATP1 and ATP2 is coincident with the subunit number of F1-α and F1-β, but that the gene dosage of ATP3 was not consistent with the subunit stoichiometry of F1F0-ATPase. By applying long PCR and gene walking analyses, we estimated that the two copies of ATP3 were approximately 20 kb apart, and we designated that which is proximal to the centromere ATP3a, while we named that which is distal ATP3b. The nucleotide sequences of the two copies of ATP3 were identical to the reported sequence in the W303-1A, W303-1B and LL20 strains, while only the DC5 strain had a single base substitution in its ATP3a. With the exception of this substitution, the other nucleotide sequences were identical to the upstream 860 bp and the downstream 150 bp. The differences between ATP3 with the single base substitution (Ser308 to Phe) and ATP3 without the substitution on the complementation of the ATP3 disruptant and on the maintenance of the mitochondrial DNA were observed, suggesting that Atp3ap and Atp3bp in the DC5 strain might have different functions. However, it should not always be necessary for yeast cells to carry different types of ATP3 because the other three strains carry the same type of ATP3. It was also demonstrated that the disruption of the ATP3 genes basically leads to a loss of wild-type mtDNA, but the stability of the mtDNA is not dependent on the ATP3 alone. Copyright © 2003 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

In this paper we report on searching for suitable reporters to monitor gene expression and protein secretion in the amylolytic yeast Schwanniomyces occidentalis. Several potential reporter and marker genes, formerly shown to be functional in other yeasts, were cloned downstream from the homologous invertase gene (INV) promoter and their activity was followed in conditions of repression and derepression of the INV promoter. However, neither β-glucuronidase nor β-lactamase nor phleomycin resistance-conferring gene, all originating from E. coli, were expressed in S. occidentalis cells to such a level to allow for monitoring of their activity. All the reporter genes tested have a higher percentage of GC (47–62%) in their DNA compared to the DNA composition of S. occidentalis genes that are more AT-rich (36% GC). The codon usage of all the reporter genes also varies from that of 16 so far sequenced S. occidentalis genes. This suggests that an appropriate composition of DNA and a codon usage similar to S. occidentalis genes might be very important parameters for an efficient expression of a heterologous gene in Schwanniomyces occidentalis. Indeed, two genes originating from Staphylococcus aureus, with an AT-content in their DNA similar to that of S. occidentalis, were functionally expressed in S. occidentalis cells. Both a phleomycin resistance-conferring gene and a chloramphenicol acetyltransferase-encoding gene thus represent suitable reporters of gene expression and protein secretion in S. occidentalis. Additionally, we show in this work that the transcription-regulating region and the signal peptide sequence of the S. occidentalis invertase gene were efficient to direct gene expression and subsequent protein secretion in Saccharomyces cerevisiae. Copyright © 2003 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Sup35 and Sup45 are essential protein components of the Saccharomyces cerevisiae translation termination factor. Yeast cells harbouring the [ PSI+] prion form of Sup35 have impaired stop codon recognition (nonsense suppression). It has long been known that the [ PSI+] prion is not stably transmitted to daughter cells when yeast are grown in the presence of m M concentrations of guanidine hydrochloride (GuHCl). In this paper, Mendelian suppressor mutations whose phenotypes are likewise hidden during growth in the presence of millimolar GuHCl are described. Such GuHCl-remedial Mendelian suppressors were selected under conditions where [ PSI+] appearance was limiting, and were caused by missense mutations in SUP35 or SUP45. Clearly, antisuppression caused by growth in the presence of GuHCl is not sufficient to distinguish missense mutations in SUP35 or SUP45, from [ PSI+]. However, the Mendelian and prion suppressors can be distinguished by subsequent growth in the absence of GuHCl, where only the nonsense suppression caused by the [ PSI+] prion remains cured. Recent reports indicate that GuHCl blocks the inheritance of [ PSI+] by directly inhibiting the activity of the protein remodelling factor Hsp104, which is required for the transmission of [ PSI+] from mother to daughter cells. However, the nonsense suppressor activity caused by the GuHCl-remedial sup35 or sup45 suppressors does not require Hsp104. Thus, GuHCl must antisuppress the sup35 and sup45 mutations via an in vivo target distinct from Hsp104. Copyright © 2003 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

The gene encoding the copper-zinc superoxide dismutase enzyme (SODC or Cu,Zn-SOD) has been cloned from several species of higher eukaryotes, but superoxide dismutase genes from moulds and yeast have not been studied extensively. Only 15 nucleotide sequences have been reported in the SwissProt, EMBL and GenBank data libraries. In general the presence of Cu,Zn-SOD in cytosol, as well as Mn-SOD in the mitochondrial matrix of yeast, has been accepted. The absence of Cu,Zn-SOD in a pigmented yeast has been accepted as a general rule. Some authors suggest that the absence of Cu,Zn-SOD in pigmented yeast is complemented by the presence of carotenoproteins that act as an extra mitochondrial antioxidant. In this report, we found that the absence of SODC is not a rule for pigmented yeast: Udeniomyces puniceus expresses an active SODC which responds to Cu2+ induction, as has been reported previously for non-pigmented yeast. The encoding region of the sod1 gene was cloned from three species of pigmented marine yeast thorough genomic DNA PCR amplification. Fragments of 485–487 nucleotides were obtained, which contain information for theoretical products of 153–154 amino acids. In Rhodotorula mucilaginosa the deduced amino acid sequence shows that insertion of three bases (C112, A149 and C166) generates a stop codon at position 123 (TGA). For Rhodotorula graminis a single change (T for A) generates a stop codon at position 298. For both species, this non-transcription of encoding sequence correlates with the absence of peptides or active proteins in cell homogenates. For U. puniceus, the cloned nucleotide sequence contains all necessary information to produce a functional protein, which correlates with activity detected in cell homogenates, both under normal conditions and by copper induction experiments. Finally, we clearly showed that the key factor in protection against oxidative stress on pigmented yeast is related not only to the presence of protective pigments but also to their amounts and spectra, as well as the presence and activity of SODC. The sod1 sequences reported in this paper have been deposited in the NCBI public data library under Accession Nos AY064723, AY064724 and AF301901. Copyright © 2003 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Ty3/gypsy retrotransposons are a widespread group of eukaryote mobile genetic elements. They are similar in structure to, and may be ancestors of, the vertebrate retroviruses. Here we describe the first Ty3/gypsy retrotransposons from the pathogenic yeasts Candida albicans and Candida dubliniensis, which we refer to as Tca3 and Tcd3, respectively. Tca3 was first identified in a variety of strains as an element lacking a large part of its coding region. Comparative analyses between C. albicans and C. dubliniensis allowed us to identify the closely related full-length Tcd3 element, and, subsequently, the full-length Tca3 elements. The full-length versions of Tca3 and Tcd3 are broadly similar in structure to other Ty3/gypsy elements, but have several features of special interest, e.g. both elements appear to have a novel mechanism for priming minus-strand DNA synthesis, probably involving conserved secondary structures adjacent to the 5' LTRs. Also, while closely related to each other, the two elements appear to be fairly distantly related to other known Ty3/gypsy-like elements. Finally, the occurrence of the internally deleted forms of Tca3 in many strains raises interesting questions concerning the evolution of these transposable elements in Candida and the evolution of Candida itself. The sequences reported in this paper have been assigned GenBank Accession Nos AF499463, AF499464 and AF510498. Copyright © 2003 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]