New applications for Schizosaccharomyces pombe in the alcoholic fermentation of red wines

The fermentation of grape must using non‐Saccharomyces yeasts with particular metabolic and biochemical properties is of growing interest. In the present work, red grape must was fermented using four strains of Schizosaccharomyces pombe (935, 936, 938 and 2139), Saccharomyces cerevisiae 7VA and Saccharomyces uvarum S6U, and comparisons were made over the fermentation period in terms of must sugar (glucose + fructose), malic acid, acetic acid, ammonia, primary amino nitrogen, lactic acid, urea (a possible fermentation activator or precursor of other metabolites) and pyruvic acid (a molecule affecting vitisin formation and therefore colour stability) concentration. The colour intensity of the fermenting musts was also recorded. The Schizosaccharomyces strains consumed less primary amino nitrogen and produced less urea and more pyruvic acid than other Saccharomyces species. Further, three of the four Schizosaccharomyces strains completed the breakdown of malic acid by day 4 of fermentation. The main negative effect of the use of Schizosaccharomyces was strong acetic acid production. The Schizosaccharomyces strains that produced most pyruvic acid (938 and 936) were associated with better ‘wine’ colour than the remaining yeasts. The studied Schizosaccharomyces could therefore be of oenological interest.     

Physiological and biochemical analysis of cytochrome P-450 in the yeast Schizosaccharomyces pombe

The yeast Schizosacchromyces pombe has been shown to contain a microsomal cytochrome P-450 (cyt. P-450) inducible under conditions of glucose repression. Under these conditions the enzyme has maximal expression of 0.43 nmol g^?1 wet wt at the end of the exponential phase of growth. Substrate and inhibitor affinity was examined using studies of spectral changes on binding and revealed a type II spectrum with ketoconazole (K_s = 23 μM ) and a type I spectrum with benzo(a)pyrene (K_s = 77 μM ). A K_m of 112 μM was found in the aryl hydrocarbon hydroxylas assay. These properties show broad comparability with the cyt. P-450 of Saccharomyces cerevisiae.     

Architectural features of pre-mRNA introns in the fission yeast Schizosaccharomyces pombe.

The architectural features of 73 introns found in 36 genes of the fission yeast Schizosaccharomyces pombe have been compiled and tabulated. The introns from S. pombe can be grouped into two size classes. Intron features are discussed in comparison to intron features of Saccharomyces cerevisiae and other eukaryotes. The results indicate that S. pombe displays quite different architectural features than the budding yeast S. cerevisiae. However, particularly in the 3' region, S. pombe introns also appear to differ from mammalian introns.     

Homologous chromosome pairing in Schizosaccharomyces pombe

Homologous chromosome pairing is a central feature of meiosis I, contributing to the correct segregation of chromosomes during meiosis. The fission yeast, Schizosaccharomyces pombe, has been widely used to study meiotic chromosome dynamics, partly because studies in this yeast are simplified due to the lack of post-pairing synaptic structures. Chromosome pairing in Sz. pombe occurs differentially throughout the genome. Telomeres cluster at the spindle pole body (SPB) at the onset of meiosis, imposing a spatial restriction on pairing events. Subsequently, centromeres dissociate from the SPB and pair in a recombination- and heterochromatin (Swi6)-independent fashion. Pairing of telomere distal regions occurs during meiotic prophase, concomitant with a dynamic association/dissociation of homologous regions, with interhomologue associations becoming increasingly stable. The stabilization of paired regions is enhanced by factors required for the initiation of meiotic recombination, suggesting that recombination stabilizes paired regions. However, substantial pairing is initiated in the absence of recombination; this is dependent upon another factor, the conserved Meu13 protein, demonstrating that recombination is not required for initial pairing interactions. During meiotic prophase Sz. pombe exhibits a pronounced dynein-dependent nuclear oscillation, which drives the pairing of centromeric and interstitial regions. Dynein is also required for the significant levels of achiasmate reductional segregation observed in Sz. pombe, possibly implicating the centromere-associated pairing with achiasmate homologue segregation. Whilst Sz. pombe does not form discernable synaptic structures continuously along the meiotic chromosomes, it does form proteinacious, meiosis-specific, linear structures (linear elements). However, the role, if any, of these structures in mediating homologue pairing is unknown.     

Metabolism of phospholipids in the yeast Schizosaccharomyces pombe

The fission yeast Schizosaccharomyces pombe is an important model organism for the study of fundamental questions in eukaryotic cell and molecular biology. A plethora of cellular processes are membrane associated and/or dependent on the proper functioning of cellular membranes. Phospholipids are not only the basic building blocks of cellular membranes; they also serve as precursors to numerous signaling molecules. In this review, we describe the biosynthetic pathways leading to major S. pombe phospholipids, how these pathways are regulated, and what is known about degradation and turnover of fission yeast phospholipids. This review also addresses the synthesis, regulation and the role of water-soluble phospholipid precursors. The last chapter of the review is devoted to the use of S. pombe for the biotechnological production of value-added lipid molecules.     

Transport of L-leucine in the fission yeast Schizosaccharomyces pombe

Transport of L -leucine into Schizosaccharomyces pombe cells from the stationary phase of growth (after preincubation for 60 min with 1% glucose) proceeds uphill, practically unidirectionally, and is mediated by at least two systems: a high-affinity system with a K_T of 0·045 mmol 1^?1 and J_max of 3·3 nmol min^?1 (mg dry weight)^?1 and a low-affinity system with a K_T of 1·25 mmol 1^?1 and J_max of 16·0 nmol min^?1 (mg dry weight)^?1. The high-affinity system has a pH optimum at 3.2, the accumulation ratio is highest at a cell density of 2–4 mg dry weight per ml and decreases with increasing leucine concentration. Transport of leucine by the high-affinity system is strongly inhibited by proton conductors, ammonium ions and by most amino acids, but only L -phenylalanine, L -isoleucine, L -valine and L -cysteine behave as fully competitive inhibitors. Systems of L -leucine transport in S. pombe are not constitutive. Transport activity appears only after preincubation of cells with a suitable source of energy. If cycloheximide is added during preincubation with glucose, no transport systems for leucine are synthesized. After removal of glucose, the activity of transport systems decays with a half-time of about 20 min. The presence of cyclic AMP increases the initial rate of leucine uptake only in cells preincubated with glucose and in the absence of cycloheximide.     

Isolation and characterization of Schizosaccharomyces pombe fragile mutants

Three Schizosaccharomyces pombe fragile mutants requiring the presence of an osmotic stabilizer to grow, that lyse when transferred into hypotonic solutions and that secrete to the extracellular medium more protein than the parental strain were isolated. In the three mutants, the fragile phenotype segregated in a Mendelian fashion, indicating a single chromosomal gene mutation, and behaved as a recessive character. By complementation analysis, the three fragile mutants fell in a single complementation group, defining the same gene (SRB1). Mutations of this gene are responsible for alterations in the cells such as fragile character, increase in the cell wall porosity, changes in the cell morphology and floc-forming ability. The study of the three srb1 alleles indicated that the degree of these alterations is proportional to a significant decrease in the galactomannan fraction of the mutants cell wall. The data presented in this report suggest that the product of the SRB1 gene is critical for the maintenance of the integrity and structure of Sz. pombe cell wall.     

Sterility (ste) genes of Schizosaccharomyces pombe

In previous experiments of Girgsdies (1982), eight sterile (ste) mutants of Schizosaccharomyces pombe did not sporulate when fused with h⁺ or h^? protoplasts. We succeeded in achieving sporulation with these mutants. Two hitherto unknown ste genes, ste7 and ste8, were found.     

Replicative ageing in the fission yeast Schizosaccharomyces pombe.

Saccharomyces cerevisiae has been widely used as a model organism in studies of replicative ageing and senescence. The relevance of these studies to ageing in other organisms has, however, been questioned, since this yeast divides by budding rather than fission, the more common pattern in higher organisms. Here we report that, contrary to popular belief, the fission yeast Schizosaccharomyces pombe also undergoes replicative senescence and in a manner superficially analogous to budding yeast. These experiments provide the first evidence of age asymmetry in cell fission and are consistent with the hypothesis of Jazwinski, that asymmetric division underlies culture immortality. Given their evolutionary divergence, comparison of the ageing determinants in fission and budding yeasts may help identify common mechanisms of the ageing process.     

Double-strand break repair and homologous recombination in Schizosaccharomyces pombe

The study of double-strand break repair and homologous recombination in Saccharomyces cerevisiae meiosis has provided important information about the mechanisms involved. However, it has become clear that the resulting recombination models are only partially applicable to repair in mitotic cells, where crossover formation is suppressed. In recent years our understanding of double-strand break repair and homologous recombination in Schizosaccharomyces pombe has increased significantly, and the identification of novel pathways and genes with homologues in higher eukaryotes has increased its value as a model organism for double-strand break repair. In this review we will focus on the involvement of homologous recombination and repair in different aspects of genome stability in Sz. pombe meiosis, replication and telomere maintenance. We will also discuss anti-recombination pathways (that suppress crossover formation), non-homologous end-joining, single-strand annealing and factors that influence the choice and prevalence of the different repair pathways in Sz. pombe.     

Enhanced deacidification activity in Schizosaccharomyces pombe by genome shuffling

A problem frequently occurring in making some kinds of wines, particularly Vitis quinquangularis Rehd wine, is the presence of malic acid at high concentrations, which is detrimental to the quality of wines. Thus, there is a need of the ways for effectively reducing the malic acid levels in wine. This study aimed to generate shuffled fusants of Schizosaccharomyces pombe with enhanced deacidification activity for reducing the excessive malic acid content in wine. Sz. pombe CGMCC 2.1628 was used as the original strain. The starting mutant population was generated by UV treatment. The mutants with higher deacidification activity were selected and subjected to recursive protoplast fusion. The resulting fusants were screened by using the indicator of malic acid concentration of fermentation supernatants on 96‐well microtitre plates, measured with bromocresol green. After three rounds of genome shuffling, the best‐performing fusant, named GS3‐1, was obtained. Its deacidification activity (consumed 4.78 g/l malic acid within 10 days) was increased by 225.2% as compared to that of original strain. In the Vitis quinquangularis Rehd wine fermentation test, GS3‐1 consumed 4.0 g/l malic acid during the whole cycle of fermentation, providing up to 185.7% improvement in malic acid consumption compared with that of the original strain. This study shows that GS3‐1 has great potential for improving the quality of Vitis quinquangularis Rehd wine. Copyright © 2014 John Wiley & Sons, Ltd.     

利用粟酒裂殖酵母生长法定量测定肌醇浓度

利用本实验室分离得到的一株天然的肌醇合成缺陷型的粟酒裂殖酵母 (Schizosaccha romycespombe)特异地依赖外部肌醇生长的特性 ,研究了该菌株在含有不同浓度肌醇的合成培养基中的生长情况 ,发现在一定的肌醇浓度范围内 ,菌株的生物量随肌醇浓度增大而增大 ,且呈线性关系 ,由此建立了一套微生物法定量测定肌醇浓度的方法。我们发现在固定的培养温度、振荡速度等外部条件下 ,当接种量OD60 0 为 1 0、葡萄糖浓度为 4 0 %时 ,在肌醇浓度 0～ 5mg/L范围内 ,培养 52h后 ,肌醇测定标准曲线最为理想 ,R值为 0 9973。应用该方法对标准肌醇样品的测定得到了较好的结果     

The mei3 region of the Schizosaccharomyces pombe genome

Expression of the mei3 gene is sufficient to induce meiosis in the fission yeast Schizosaccharomyces pombe. The mei3 gene is located 0.64 Mb from the telomere of the left arm of Sz. pombe chromosome II. We have sequenced and analysed 107 kb of DNA from the mei3 genomic region. The sequence includes 14 known genes (bag1-B, csh3, dps1, gpt1, mei3, mfm3, pac1, prp31, rpl38-1, rpn3, rti1, spa1, spm1 and ubc4) and 26 other open reading frames (ORFs) longer than 100 codons: a density of one protein-coding gene per 2.7 kb. Twenty-one of the 40 ORFs (53%) have introns. In addition there is one lone Tf1 transposon long terminal repeat (LTR), tRNA(Trp) and tRNA(Ser) genes and a 5S rRNA gene. 14 of the novel ORFs show sequence similarities which suggest functions of their products, including a coatomer alpha-subunit, a catechol O-methyltransferase, protein kinase, asparagine synthetase, zinc metalloprotease, acetyltransferase, phosphatidylinositol 4-kinase, inositol polyphosphate phosphatase, GTPase-activating protein, permease, pre-mRNA splicing factor, 20S proteasome component and a thioredoxin-like protein. One predicted protein has similarity to the human Cockayne syndrome protein CSA and one with human GTPase XPA binding protein XAB1. Three ORFs are likely to code for proteins because they have sequence similarity with hypothetical proteins, three encode predicted coiled-coil proteins and four are sequence orphans.     

Interpretations of ultraviolet absorption in white wines

Methods are described for the general analysis of total ultraviolet (u.v.) absorbing constituents of white wines. An elution profile, which is characteristic of the particular wine, is obtained by continuous monitoring of effluent from a dextran gel column at 280 nm. Surprisingly, about half of the total u.v. absorbance in the Riesling wine examined was found to be due to non-phenolic wine constituents, which are separately eluted in the gel column analysis. These are nucleotidic materials, to which little attention has previously been given in oenology. Data relating to the protein-tannin fraction, and to nucleotidic and phenolic constituents, are directly obtained by recording the wine elution profiles at 265, 280 and 320 nm. The interpretation of the elution curves and the significance and general utility of this new analytical method are discussed.     

Biotransformation of steroids by the fission yeast Schizosaccharomyces pombe

The fungal biotransformation of steroids is of applied interest due to the economic importance of such stereo‐ and regiospecific reactions and also in the context of ergosterol pathway engineering to produce vitamin D and steroidal products. In Schizosaccharomyces pombe no steroid hydroxylation as is found in filamentous fungi was observed, but a cytosolic NAD(H)/NADP(H)‐dependent hydroxysteroid dehydrogenase activity was identified. Progesterone was reduced at the Δ⁴ double bond (in vivo only) as well as at the C‐3 and C‐20 keto groups. Testosterone and 4‐androstene‐3,17‐dione were interconverted and 5α‐pregnane‐3,20‐dione and 5β‐pregnane‐3,20‐dione were reduced to 3‐hydroxy products. The reactions were sometimes reversible and showed regio‐ and stereo specificity. In S. pombe more than one steroid dehydrogenase homologue is likely to occur, as has been observed in Saccharomyces cerevisiae. Our findings indicate that genes encoding soluble proteins should be examined as candidates for actual steroid dehydrogenase activity. Copyright © 1999 John Wiley & Sons, Ltd.     

Purification and characterization of aminopeptidase yspI from Schizosaccharomyces pombe

Aminopeptidase yspI was purified to apparent homogeneity from the fission yeast Schizosaccharomyces pombe. The molecular mass of the native enzyme was estimated to be 184 kDa by gel filtration chromatography. A value of 92 kDa was calculated after sodium dodecyl sulfate–polyacrylamide gel electrophoresis. The enzyme is thus a dimer with two identical subunits. Optimum pH for cleavage of synthetic aminoacyl-4-nitroanilides is 7·0. Mercury ions, EDTA and chloroquine were found to be potent inhibitors of aminopeptidase yspI activity. Substrate specificity studies indicate that the purified enzyme cleaves L-lysine-4-nitroanilide with high efficiency.     

The cyclophilin repertoire of the fission yeast Schizosaccharomyces pombe

The cyclophilin repertoire of the fission yeast Schizosaccharomyces pombe is comprised of nine members that are distributed over all three of its chromosomes and range from small single-domain to large multi-domain proteins. Each cyclophilin possesses only a single prolyl-isomerase domain, and these vary in their degree of consensus, including at positions that are likely to affect their drug-binding ability and catalytic activity. The additional identified motifs are involved in putative protein or RNA interactions, while a novel domain that is specific to SpCyp7 and its orthologues may have functions that include an interaction with hnRNPs. The Sz. pombe cyclophilins are found throughout the cell but appear to be absent from the mitochondria, which is unique among the characterized eukaryotic repertoires. SpCyp5, SpCyp6 and SpCyp8 have exhibited significant upregulation of their expression during the meiotic cycle and SpCyp5 has exhibited significant upregulation of its expression during heat stress. All nine have identified members in the repertoires of H. sapiens, D. melanogaster and A. thaliana. However, only three identified members in the cyclophilin repertoire of S. cerevisiae with SpCyp7 identifying a fourth protein that is not a member of the recognized repertoire due to its possession of a degenerate prolyl-isomerase domain. The cyclophilin repertoire of Sz. pombe therefore represents a better model group for the study of cyclophilin function in the higher eukaryotes.     

Characterization of a branched-chain amino-acid aminotransferase from Schizosaccharomyces pombe

The Saccharomyces cerevisiae genes for the cytosolic and mitochondrial branched-chain amino-acid aminotransferases (BCAT) were isolated recently. These genes show significant homology to mammalian ECA39, originally isolated as a gene regulated by the c-myc oncogene. We now report the isolation of the Schizosaccharomyces pombe eca39/BCAT gene. The S. pombe protein shows 47–52% identity to other eukaryotic BCAT proteins isolated from S. cerevisiae, nematode, mouse and man. A genetic growth assay for BCAT activity was established using an S. cerevisiae strain disrupted in both BCAT isoenzymes. Consequently, the activity of the S. pombe BCAT was demonstrated by genetic and biochemical means. Possible applications of BCAT-encoding genes as selection markers in yeast transformation are proposed. The sequence has been deposited in the GenBank data library under Accession Number U88029. © 1998 John Wiley & Sons, Ltd.     

Volatile components of Zalema white wines

The volatile composition of young white wines from Vitis vinifera cv. Zalema, an autochthonous grape variety in Huelva (southern Spain), has been studied by gas chromatography-olfactometry (GC-O) and techniques of quantitative analysis. This is the first time that an olfactometric analysis has been reported in wines made from this grape variety. The quantitative chemical study has shown 71 volatile compounds, of which 23 were in concentrations above their thresholds. On the basis of the odour activity values (OAVs), the most potent odorants were fermentative compounds, mainly fatty acids and their ethyl esters. Two norisoprenoids, β-damascenone and β-ionone, two alcohols (isoamyl alcohol and β-phenylethanol), three volatile thiols, 4-mercapto-4-methyl-2-pentanone, 3-mercaptohexyl acetate and 3-mercapto-1-hexanol, and two carbonyl compounds (acetaldehyde and phenylacetaldehyde) also exhibited OAVs > 1. The GC-O study corroborated these results, showing that five esters (isoamyl acetate, ethyl hexanoate, ethyl butyrate, ethyl isovalerate and ethyl octanoate), isoamyl alcohol and β-damascenone can be considered as the most powerful odorants of Zalema wines.     

Characterization of two fructosyl-amino acid oxidase homologs of Schizosaccharomyces pombe

Two putative fructosyl-amino acid oxidase genes, FAP1 and FAP2, found in the Schizosaccharomyces pombe genome were cloned and expressed. Both of the gene products (Fap1 and Fap2) were flavoproteins and have no activity for fructosyl-amino acids. It was suggested that Fap1 and Fap2 are an L-pipecolic acid oxidase and L-saccharopine oxidase, respectively.