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1.
Proc Natl Acad Sci U S A ; 116(39): 19415-19420, 2019 09 24.
Artículo en Inglés | MEDLINE | ID: mdl-31467169

RESUMEN

Biobased C4-dicarboxylic acids are attractive sustainable precursors for polymers and other materials. Commercial scale production of these acids at high titers requires efficient secretion by cell factories. In this study, we characterized 7 dicarboxylic acid transporters in Xenopus oocytes and in Saccharomyces cerevisiae engineered for dicarboxylic acid production. Among the tested transporters, the Mae1(p) from Schizosaccharomyces pombe had the highest activity toward succinic, malic, and fumaric acids and resulted in 3-, 8-, and 5-fold titer increases, respectively, in S. cerevisiae, while not affecting growth, which was in contrast to the tested transporters from the tellurite-resistance/dicarboxylate transporter (TDT) family or the Na+ coupled divalent anion-sodium symporter family. Similar to SpMae1(p), its homolog in Aspergillus carbonarius, AcDct(p), increased the malate titer 12-fold without affecting the growth. Phylogenetic and protein motif analyses mapped SpMae1(p) and AcDct(p) into the voltage-dependent slow-anion channel transporter (SLAC1) clade of transporters, which also include plant Slac1(p) transporters involved in stomata closure. The conserved phenylalanine residue F329 closing the transport pore of SpMae1(p) is essential for the transporter activity. The voltage-dependent SLAC1 transporters do not use proton or Na+ motive force and are, thus, less energetically expensive than the majority of other dicarboxylic acid transporters. Such transporters present a tremendous advantage for organic acid production via fermentation allowing a higher overall product yield.


Asunto(s)
Ácidos Dicarboxílicos/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Aminoácidos , Animales , Aspergillus/clasificación , Aspergillus/genética , Fermentación , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Mutación , Oocitos/metabolismo , Transportadores de Anión Orgánico/química , Transportadores de Anión Orgánico/genética , Filogenia , Conformación Proteica , Saccharomyces cerevisiae/clasificación , Schizosaccharomyces/clasificación , Schizosaccharomyces/genética , Xenopus/genética , Xenopus/metabolismo
2.
PLoS Genet ; 14(3): e1007288, 2018 03.
Artículo en Inglés | MEDLINE | ID: mdl-29596413

RESUMEN

Modification defects in the tRNA anticodon loop often impair yeast growth and cause human disease. In the budding yeast Saccharomyces cerevisiae and the phylogenetically distant fission yeast Schizosaccharomyces pombe, trm7Δ mutants grow poorly due to lack of 2'-O-methylation of C32 and G34 in the tRNAPhe anticodon loop, and lesions in the human TRM7 homolog FTSJ1 cause non-syndromic X-linked intellectual disability (NSXLID). However, it is unclear why trm7Δ mutants grow poorly. We show here that despite the fact that S. cerevisiae trm7Δ mutants had no detectable tRNAPhe charging defect in rich media, the cells constitutively activated a robust general amino acid control (GAAC) response, acting through Gcn2, which senses uncharged tRNA. Consistent with reduced available charged tRNAPhe, the trm7Δ growth defect was suppressed by spontaneous mutations in phenylalanyl-tRNA synthetase (PheRS) or in the pol III negative regulator MAF1, and by overexpression of tRNAPhe, PheRS, or EF-1A; all of these also reduced GAAC activation. Genetic analysis also demonstrated that the trm7Δ growth defect was due to the constitutive robust GAAC activation as well as to the reduced available charged tRNAPhe. Robust GAAC activation was not observed with several other anticodon loop modification mutants. Analysis of S. pombe trm7 mutants led to similar observations. S. pombe Trm7 depletion also resulted in no observable tRNAPhe charging defect and a robust GAAC response, and suppressors mapped to PheRS and reduced GAAC activation. We speculate that GAAC activation is widely conserved in trm7 mutants in eukaryotes, including metazoans, and might play a role in FTSJ1-mediated NSXLID.


Asunto(s)
Aminoácidos/metabolismo , Anticodón , ARN de Transferencia/metabolismo , Saccharomyces cerevisiae/clasificación , Saccharomyces cerevisiae/genética , Schizosaccharomyces/clasificación , Schizosaccharomyces/genética , Genes Fúngicos , Metilación , Mutación , Filogenia , Saccharomyces cerevisiae/crecimiento & desarrollo , Schizosaccharomyces/crecimiento & desarrollo
3.
FEMS Yeast Res ; 19(4)2019 06 01.
Artículo en Inglés | MEDLINE | ID: mdl-31132130

RESUMEN

Eight yeast strains that asexually reproduce by cell fission were isolated from bee bread of different solitary bees in Germany. DNA sequence analysis revealed that the strains shared the same sequence in the D1/D2 domain of the nuclear large subunit (LSU) rRNA gene with a strain that was previously isolated from a fig snack from Spain. The closest related type strain was that of Schizosaccharomyces octosporus, which showed 98.2% sequence similarity (11 substitutions) with the new strains. By clone sequence analysis of the internal transcribed spacer (ITS) region (ITS1, 5.8S rDNA, and ITS2) a total of nine different copy types were identified. The new strains differed from S. octosporus by approximately 31% in the ITS region. Sequence analysis of the RNAse P gene further supported the description of a new species. The strains isolated during this study show some phenotypic characteristics that separate them from the closest related species, S. octosporus and S. cryophilus. Since all strains showed true osmophily the name of the new species is S. osmophilus (holotype: CBS 15793T; isotype: CLIB 3267 T = NCAIM Y.02225 T, MycoBank no.: MB829586).


Asunto(s)
Abejas/microbiología , Própolis , Schizosaccharomyces/clasificación , Schizosaccharomyces/fisiología , Animales , Análisis por Conglomerados , ADN de Hongos/genética , ADN Intergénico/genética , Alemania , Técnicas de Tipificación Micológica , Filogenia , ARN Ribosómico/genética , Reproducción Asexuada , Schizosaccharomyces/aislamiento & purificación , Análisis de Secuencia de ADN
4.
Crit Rev Biochem Mol Biol ; 51(2): 86-95, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-26523839

RESUMEN

Next-generation sequencing approaches have considerably advanced our understanding of genome function and regulation. However, the knowledge of gene function and complex cellular processes remains a challenge and bottleneck in biological research. Phenomics is a rapidly emerging area, which seeks to rigorously characterize all phenotypes associated with genes or gene variants. Such high-throughput phenotyping under different conditions can be a potent approach toward gene function. The fission yeast Schizosaccharomyces pombe (S. pombe) is a proven eukaryotic model organism that is increasingly used for genomewide screens and phenomic assays. In this review, we highlight current large-scale, cell-based approaches used with S. pombe, including computational colony-growth measurements, genetic interaction screens, parallel profiling using barcodes, microscopy-based cell profiling, metabolomic methods and transposon mutagenesis. These diverse methods are starting to offer rich insights into the relationship between genotypes and phenotypes.


Asunto(s)
Genoma Viral , Fenotipo , Schizosaccharomyces/genética , Aminoácidos/metabolismo , Elementos Transponibles de ADN , Metabolómica , Modelos Biológicos , Mutagénesis , Schizosaccharomyces/clasificación
5.
Mol Cell Proteomics ; 15(10): 3090-3106, 2016 10.
Artículo en Inglés | MEDLINE | ID: mdl-27477394

RESUMEN

Protein secretion in yeast is a complex process and its efficiency depends on a variety of parameters. We performed a comparative proteome analysis of a set of Schizosaccharomyces pombe strains producing the α-glucosidase maltase in increasing amounts to investigate the overall proteomic response of the cell to the burden of protein production along the various steps of protein production and secretion. Proteome analysis of these strains, utilizing an isobaric labeling/two dimensional LC-MALDI MS approach, revealed complex changes, from chaperones and secretory transport machinery to proteins controlling transcription and translation. We also found an unexpectedly high amount of changes in enzyme levels of the central carbon metabolism and a significant up-regulation of several amino acid biosyntheses. These amino acids were partially underrepresented in the cellular protein compared with the composition of the model protein. Additional feeding of these amino acids resulted in a 1.5-fold increase in protein secretion. Membrane fluidity was identified as a second bottleneck for high-level protein secretion and addition of fluconazole to the culture caused a significant decrease in ergosterol levels, whereas protein secretion could be further increased by a factor of 2.1. In summary, we show that high level protein secretion causes global changes of protein expression levels in the cell and that precursor availability and membrane composition limit protein secretion in this yeast. In this respect, comparative proteome analysis is a powerful tool to identify targets for an efficient increase of protein production and secretion in S. pombe Data are available via ProteomeXchange with identifiers PXD002693 and PXD003016.


Asunto(s)
Proteómica/métodos , Proteínas de Schizosaccharomyces pombe/metabolismo , Schizosaccharomyces/metabolismo , Cromatografía Liquida , Regulación Fúngica de la Expresión Génica , Schizosaccharomyces/clasificación , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , alfa-Glucosidasas/metabolismo
6.
Genes Cells ; 21(6): 594-607, 2016 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-27059155

RESUMEN

In the fission yeast Schizosaccharomyces pombe (Sp), Mid1/Dmf1 plays an important role in positioning the division site by inducing formation of the contractile ring (CR). Mid1, emanating from the nucleus located in the cell center, forms a dozen of nodes in the middle cell cortex ahead of mitosis, and actin filaments and myosin II accumulated at each node interact and assemble the CR in metaphase. Curiously, in another fission yeast S. japonicus (Sj), CR formation begins after nuclear segregation in late anaphase. Here, we investigated the role of S. japonicus Mid1 during mitosis to compare the molecular mechanisms that determine the cell division site in Schizosaccharomyces. Similar to Sp Mid1, Sj Mid1 often accumulated in the nucleus of interphase cells. Moreover, Sj Mid1 localized to cortical dots with myosin II in the future division site and formed a medial ring in mitotic cells. However, S. japonicus cells without Mid1 function still carried out symmetrical binary division. Therefore, the Mid1 dependency for positional control of the cell division site is possibly different between the two species. Meanwhile, we found that Sj Mid1 enhanced CR formation, in a manner possibly similar to that by Sp Mid1.


Asunto(s)
Proteínas Fúngicas/metabolismo , Mitosis , Schizosaccharomyces/citología , Schizosaccharomyces/metabolismo , Actinas/metabolismo , Anafase , Citocinesis , Miosinas/metabolismo , Schizosaccharomyces/clasificación , Proteínas de Schizosaccharomyces pombe/metabolismo
7.
Genes Cells ; 21(6): 530-42, 2016 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-27005325

RESUMEN

Living organisms have evolved multiple sophisticated mechanisms to deal with reactive oxygen species. We constructed a collection of twelve single-gene deletion strains of the fission yeast Schizosaccharomyces pombe designed for the study of oxidative and heavy metal stress responses. This collection contains deletions of biosynthetic enzymes of glutathione (Δgcs1 and Δgsa1), phytochelatin (Δpcs2), ubiquinone (Δabc1) and ergothioneine (Δegt1), as well as catalase (Δctt1), thioredoxins (Δtrx1 and Δtrx2), Cu/Zn- and Mn- superoxide dismutases (SODs; Δsod1 and Δsod2), sulfiredoxin (Δsrx1) and sulfide-quinone oxidoreductase (Δhmt2). First, we employed metabolomic analysis to examine the mutants of the glutathione biosynthetic pathway. We found that ophthalmic acid was produced by the same enzymes as glutathione in S. pombe. The identical genetic background of the strains allowed us to assess the severity of the individual gene knockouts by treating the deletion strains with oxidative agents. Among other results, we found that glutathione deletion strains were not particularly sensitive to peroxide or superoxide, but highly sensitive to cadmium stress. Our results show the astonishing diversity in cellular adaptation mechanisms to various types of oxidative and metal stress and provide a useful tool for further research into stress responses.


Asunto(s)
Metales Pesados/toxicidad , Estrés Oxidativo , Schizosaccharomyces/fisiología , Vías Biosintéticas , Eliminación de Gen , Glutatión/genética , Oligopéptidos/biosíntesis , Schizosaccharomyces/clasificación , Schizosaccharomyces/efectos de los fármacos , Schizosaccharomyces/genética , Proteínas de Schizosaccharomyces pombe/genética , Proteínas de Schizosaccharomyces pombe/metabolismo , Estrés Fisiológico
8.
Curr Microbiol ; 74(6): 710-717, 2017 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-28342076

RESUMEN

Environmental glucose is an important regulator of biological processes, as it can launch different cell processes depending on its concentration. Thus, low glucose concentration can induce entry into quiescence, which ensures long-term viability for the cells or in other cases mycelial growth in the dimorphic species, which, in turn, provides the cells with fresh nutrients. Several genes, such as the genes of cAMP cascade, are involved in glucose sensing and response. Since this signal transduction pathway seemed to be an evolutionarily conserved process, we assumed that its genes were also conserved and preserved their functional homology. To obtain evidence, Schizosaccharomyces pombe rsv1 and its orthologous genes were investigated using in silico and experimental approaches. Our results supported that the Rsv1 zinc-finger transcription factors of Schizosaccharomyces japonicus and Schizosaccharomyces octosporus and the Candida albicans cas5p were really functional homologues of the S. pombe Rsv1. Namely, the homologous proteins were able to restore mutant phenotype of the S. pombe rsv1-deleted cells. Bioinformatic anaysis revealed that the most conserved parts of the proteins always contained the C2H2 domains and the complementation abilities of the counterpart genes were not uniform regarding the investigated features, which can be in connection with the conserved regions outside C2H2.


Asunto(s)
Candida albicans/genética , Proteínas de Unión al ADN/genética , Glucosa/metabolismo , Proteínas de Schizosaccharomyces pombe/genética , Schizosaccharomyces/genética , Inanición/genética , Factores de Transcripción/genética , Ácido Acético/farmacología , Secuencia de Aminoácidos , Cafeína/farmacología , Caspofungina , Rojo Congo/farmacología , AMP Cíclico/metabolismo , ADN de Hongos/genética , Equinocandinas/farmacología , Evolución Molecular , Eliminación de Gen , Lipopéptidos/farmacología , Schizosaccharomyces/clasificación , Schizosaccharomyces/metabolismo , Homología de Secuencia de Aminoácido , Transducción de Señal/genética
9.
Food Microbiol ; 61: 136-149, 2017 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-27697163

RESUMEN

The present work demonstrates that yeasts belonging to the Schizosaccharomyces genus release a high quantity of polysaccharides of cell wall origin starting from the onset of the alcoholic fermentation. By the end of the alcoholic fermentation, all of the Schizosaccharomyces yeast strains released a quantity of polysaccharides approximately 3-7 times higher than that released by a commercial Saccharomyces cerevisiae yeast strain under the same fermentative conditions of synthetic juice. A higher content of polysaccharide was found in media fermented by Schizosaccharomyces japonicus with respect to that of Schizosaccharomyces pombe. Some of the strains evaluated were also able to produce high levels of pyruvic acid, which has been shown to be an important compound for color stability of wine. The presence of strains with different malic acid consumption patterns along with high polysaccharide release would enable production of naturally modified wines with enhanced mouth feel and reduced acidity. The chemical analysis of the released polysaccharides demonstrated divergence between the two yeast species S. pombe and S. japonicus. A different mannose/galactose ratio and a different percentage of proteins was observed on the polysaccharides released by S. pombe as compared to S. japonicus. Analysis of the proteins released in the media revealed the presence of a glycoprotein with a molecular size around 32-33 kDa only for the species S. japonicus. Mass spectrometry analysis of carbohydrate moieties showed similar proportions among the N-glycan chains released in the media by both yeast species but differences between the two species were also observed. These observations suggest a possible role of rapid MALDI-TOF screening of N-glycans compositional fingerprint as a taxonomic tool for this genus. Polysaccharides release in the media, in particular galactomannoproteins in significant amounts, could make these yeasts particularly interesting also for the industrial production of exogenous polysaccharide preparations.


Asunto(s)
Pared Celular/química , Polisacáridos/química , Polisacáridos/metabolismo , Saccharomyces cerevisiae/metabolismo , Schizosaccharomyces/metabolismo , Fermentación , Galactosa/análogos & derivados , Mananos/biosíntesis , Glicoproteínas de Membrana/biosíntesis , Polisacáridos/análisis , Polisacáridos/aislamiento & purificación , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/clasificación , Schizosaccharomyces/química , Schizosaccharomyces/clasificación , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción , Vino/análisis , Vino/microbiología
10.
Antonie Van Leeuwenhoek ; 108(1): 97-106, 2015 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-25948336

RESUMEN

Investigation of yeast neutral lipid accumulation is important for biotechnology and also for modelling aberrant lipid metabolism in human disease. The Nile red (NR) method has been extensively utilised to determine lipid phenotypes of yeast cells via microscopic means. NR assays have been used to differentiate lipid accumulation and relative amounts of lipid in oleaginous species but have not been thoroughly validated for phenotype determination arising from genetic modification. A modified NR assay, first described by Sitepu et al. (J Microbiol Methods 91:321-328, 2012), was able to detect neutral lipid changes in Saccharomyces cerevisiae deletion mutants with sensitivity similar to more advanced methodology. We have also be able to, for the first time, successfully apply the NR assay to the well characterised fission yeast Schizosaccharomyces pombe, an increasingly important organism in biotechnology. The described NR fluorescence assay is suitable for increased throughput and rapid screening of genetically modified strains in both the biotechnology industry and for modelling ectopic lipid production for a variety of human diseases. This ultimately negates the need for labour intensive and time consuming lipid analyses of samples that may not yield a desirable lipid phenotype, whilst genetic modifications impacting significantly on the cellular lipid phenotype can be further promoted for more in depth analyses.


Asunto(s)
Lípidos/análisis , Oxazinas/metabolismo , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/clasificación , Schizosaccharomyces/química , Schizosaccharomyces/clasificación , Coloración y Etiquetado/métodos , Fluorescencia , Metabolismo de los Lípidos , Tamizaje Masivo/métodos , Fenotipo , Saccharomyces cerevisiae/metabolismo , Schizosaccharomyces/metabolismo
11.
Fungal Genet Biol ; 49(3): 235-49, 2012 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-22300943

RESUMEN

The fission yeasts (Schizosaccharomyces) representing a highly divergent phylogenetic branch of Fungi evolved from filamentous ancestors by gradual transition from mycelial growth to yeast morphology. For the transition, a mechanism had been developed that separates the sister cells after the completion of cytokinesis. Numerous components of the separation mechanism have been characterised in Schizosaccharomycespombe, including the zinc-finger transcription factor Ace2p and the fork-head transcription factor Sep1p. Here we show that both regulators have regions conserved within the genus. The most conserved parts contain the DNA-binding domains whose amino-acid sequences perfectly reflect the phylogenetic positions of the species. The less conserved parts of the proteins contain sequence blocks specific for the whole genus or only for the species propagating predominantly or exclusively as yeasts. Inactivation of either gene in the dimorphic species Schizosaccharomycesjaponicus abolished cell separation in the yeast phase conferring hypha-like morphology but did not change the growth pattern to unipolar and did not cause extensive polar vacuolation characteristic of the true mycelium. Neither mutation affected the mycelial phase, but both mutations hampered the hyphal fragmentation at the mycelium-to-yeast transition. Ace2p(Sj) acts downstream of Sep1p(Sj) and regulates the orthologues of the Ace2p-dependent S.pombe genes agn1(+) (1,3-alpha-glucanase) and eng1(+) (1,3-beta-glucanase) but does not regulate the orthologue of cfh4(+) (chitin synthase regulatory factor). These results and the complementation of the cell separation defects of the ace2(-) and sep1(-) mutations of S.pombe by heterologously expressed ace2(Sj) and sep1(Sj) indicate that the cell separation mechanism is conserved in the Schizosaccharomyces genus.


Asunto(s)
División Celular , Regulación Fúngica de la Expresión Génica , Proteínas de Schizosaccharomyces pombe/genética , Schizosaccharomyces/citología , Schizosaccharomyces/genética , Factores de Transcripción/genética , Secuencia de Aminoácidos , Secuencia Conservada , Datos de Secuencia Molecular , Filogenia , Estructura Terciaria de Proteína , Schizosaccharomyces/clasificación , Schizosaccharomyces/metabolismo , Proteínas de Schizosaccharomyces pombe/química , Proteínas de Schizosaccharomyces pombe/metabolismo , Factores de Transcripción/química , Factores de Transcripción/metabolismo
12.
Mol Biol Evol ; 26(1): 27-34, 2009 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-18922765

RESUMEN

Several morphologically dissimilar ascomycete fungi including Schizosaccharomyces, Taphrina, Saitoella, Pneumocystis, and Neolecta have been grouped into the taxon Taphrinomycotina (Archiascomycota or Archiascomycotina), originally based on rRNA phylogeny. These analyses lack statistically significant support for the monophyly of this grouping, and although confirmed by more recent multigene analyses, this topology is contradicted by mitochondrial phylogenies. To resolve this inconsistency, we have assembled phylogenomic mitochondrial and nuclear data sets from four distantly related taphrinomycotina taxa: Schizosaccharomyces pombe, Pneumocystis carinii, Saitoella complicata, and Taphrina deformans. Our phylogenomic analyses based on nuclear data (113 proteins) conclusively support the monophyly of Taphrinomycotina, diverging as a sister group to Saccharomycotina + Pezizomycotina. However, despite the improved taxon sampling, Taphrinomycotina continue to be paraphyletic with the mitochondrial data set (13 proteins): Schizosaccharomyces species associate with budding yeasts (Saccharomycotina) and the other Taphrinomycotina group as a sister group to Saccharomycotina + Pezizomycotina. Yet, as Schizosaccharomyces and Saccharomycotina species are fast evolving, the mitochondrial phylogeny may be influenced by a long-branch attraction (LBA) artifact. After removal of fast-evolving sequence positions from the mitochondrial data set, we recover the monophyly of Taphrinomycotina. Our combined results suggest that Taphrinomycotina is a legitimate taxon, that this group of species diverges as a sister group to Saccharomycotina + Pezizomycotina, and that phylogenetic positioning of yeasts and fission yeasts with mitochondrial data is plagued by a strong LBA artifact.


Asunto(s)
Schizosaccharomyces/clasificación , Schizosaccharomyces/genética , Ascomicetos/clasificación , Ascomicetos/genética , ADN Complementario/genética , ADN de Hongos/genética , Etiquetas de Secuencia Expresada , Proteínas Fúngicas/genética , Proteínas Mitocondriales/genética , Proteínas Nucleares/genética , Filogenia , Schizosaccharomyces/citología
13.
FEMS Yeast Res ; 10(6): 779-86, 2010 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-20618870

RESUMEN

The genus Schizosaccharomyces is presently comprised of three species, namely Schizosaccharomyces pombe, Schizosaccharomyces octosporus and Schizosaccharomyces japonicus. Here, we describe a hitherto unknown species, Schizosaccharomyces cryophilus, named for its preference for growth at lower temperatures than the other fission yeast species. Although morphologically similar to S. octosporus, analysis of several rapidly evolving sequences, including the D1/D2 divergent domain of the large subunit (LSU) rRNA gene, the RNA subunit of RNAse P and the internal transcribed spacer elements, revealed significant divergence from any previously characterized Schizosaccharomyces strain. Based on phylogenetic analysis of the D1/D2 domain of the LSU rRNA gene, S. octosporus is the closest known relative of S. cryophilus, with the sequences of the two species differing by 25 nucleotide substitutions (>4%). Sequencing of the S. cryophilus genome and phylogenetic analysis of all 1 : 1 protein orthologs confirmed this observation, and together with morphological and physiological characterization, supports the assignment of S. cryophilus as a new species within the genus Schizosaccharomyces. The type strain of the new species is NRRL Y-48691(T) (=NBRC 106824(T)=CBS 11777(T)).


Asunto(s)
Schizosaccharomyces/clasificación , Análisis por Conglomerados , Frío , ADN de Hongos/química , ADN de Hongos/genética , ADN Ribosómico/química , ADN Ribosómico/genética , ADN Espaciador Ribosómico/química , ADN Espaciador Ribosómico/genética , Proteínas Fúngicas/genética , Genes de ARNr , Genoma Fúngico , Datos de Secuencia Molecular , Filogenia , ARN de Hongos/genética , ARN Ribosómico/genética , Ribonucleasa P/genética , Schizosaccharomyces/citología , Schizosaccharomyces/genética , Schizosaccharomyces/fisiología , Análisis de Secuencia de ADN
14.
Genome Biol Evol ; 11(8): 2312-2329, 2019 08 01.
Artículo en Inglés | MEDLINE | ID: mdl-31364709

RESUMEN

The fission yeast Schizosaccharomyces pombe is an important model organism, but its natural diversity and evolutionary history remain under-studied. In particular, the population genomics of the S. pombe mitochondrial genome (mitogenome) has not been thoroughly investigated. Here, we assembled the complete circular-mapping mitogenomes of 192 S. pombe isolates de novo, and found that these mitogenomes belong to 69 nonidentical sequence types ranging from 17,618 to 26,910 bp in length. Using the assembled mitogenomes, we identified 20 errors in the reference mitogenome and discovered two previously unknown mitochondrial introns. Analyzing sequence diversity of these 69 types of mitogenomes revealed two highly distinct clades, with only three mitogenomes exhibiting signs of inter-clade recombination. This diversity pattern suggests that currently available S. pombe isolates descend from two long-separated ancestral lineages. This conclusion is corroborated by the diversity pattern of the recombination-repressed K-region located between donor mating-type loci mat2 and mat3 in the nuclear genome. We estimated that the two ancestral S. pombe lineages diverged about 31 million generations ago. These findings shed new light on the evolution of S. pombe and the data sets generated in this study will facilitate future research on genome evolution.


Asunto(s)
Evolución Molecular , Variación Genética , Genoma Mitocondrial , Metagenómica , Recombinación Genética , Schizosaccharomyces/clasificación , Schizosaccharomyces/genética , Filogenia
15.
Food Res Int ; 126: 108582, 2019 12.
Artículo en Inglés | MEDLINE | ID: mdl-31732056

RESUMEN

Vitis davidii Föex is widely planted in South China as a potential wine making grape. In this study, yeast communities during the spontaneous fermentations of two varieties of Vitis davidii Föex in Guizhou, China were investigated. Hanseniaspora uvarum, Pichia terricola, Saccharomyces cerevisiae/S. mikatae, and Schizosaccharomyces japonicus were the four main yeast species detected by culture-dependent and high-throughput sequencing approaches. However, the composition of minor yeast species was quite different as revealed by the two approaches. Ten yeast species including Debaryomyces hansenii, Rhodotorula mucilaginosa, Sporidiobolus paraoseus, Starmerella bacillaris, Zygoascus meyerae, etc. were detected by culture-dependent approaches, whereas the other five species were found by high-throughput sequencing analysis. S. japonicus was widely found during spontaneous fermentations and its concentrations were mostly higher than that of S. cerevisiae. The difference in grape varieties and fermentation conditions contributed to yeast diversity. The results of this study provide basic information on indigenous yeast diversity from Vitis davidii Föex in Guizhou, which would help to exploit the potential of non-Saccharomyces yeast species with good oenological attributes.


Asunto(s)
Saccharomyces cerevisiae/metabolismo , Vitis/química , Vitis/microbiología , China , ADN de Hongos/aislamiento & purificación , Fermentación , Microbiología de Alimentos , Hanseniaspora/clasificación , Hanseniaspora/metabolismo , Secuenciación de Nucleótidos de Alto Rendimiento , Pichia/clasificación , Pichia/metabolismo , Saccharomyces cerevisiae/clasificación , Saccharomycetales/clasificación , Saccharomycetales/metabolismo , Schizosaccharomyces/clasificación , Schizosaccharomyces/metabolismo , Análisis de Secuencia de ADN , Vino/análisis , Vino/microbiología
16.
Nat Protoc ; 14(2): 415-440, 2019 02.
Artículo en Inglés | MEDLINE | ID: mdl-30635653

RESUMEN

The construction of genome-wide mutant collections has enabled high-throughput, high-dimensional quantitative characterization of gene and chemical function, particularly via genetic and chemical-genetic interaction experiments. As the throughput of such experiments increases with improvements in sequencing technology and sample multiplexing, appropriate tools must be developed to handle the large volume of data produced. Here, we describe how to apply our approach to high-throughput, fitness-based profiling of pooled mutant yeast collections using the BEAN-counter software pipeline (Barcoded Experiment Analysis for Next-generation sequencing) for analysis. The software has also successfully processed data from Schizosaccharomyces pombe, Escherichia coli, and Zymomonas mobilis mutant collections. We provide general recommendations for the design of large-scale, multiplexed barcode sequencing experiments. The procedure outlined here was used to score interactions for ~4 million chemical-by-mutant combinations in our recently published chemical-genetic interaction screen of nearly 14,000 chemical compounds across seven diverse compound collections. Here we selected a representative subset of these data on which to demonstrate our analysis pipeline. BEAN-counter is open source, written in Python, and freely available for academic use. Users should be proficient at the command line; advanced users who wish to analyze larger datasets with hundreds or more conditions should also be familiar with concepts in analysis of high-throughput biological data. BEAN-counter encapsulates the knowledge we have accumulated from, and successfully applied to, our multiplexed, pooled barcode sequencing experiments. This protocol will be useful to those interested in generating their own high-dimensional, quantitative characterizations of gene or chemical function in a high-throughput manner.


Asunto(s)
Interacción Gen-Ambiente , Genoma Bacteriano , Genoma Fúngico , Saccharomyces cerevisiae/genética , Bibliotecas de Moléculas Pequeñas/farmacología , Programas Informáticos , Código de Barras del ADN Taxonómico/métodos , ADN Bacteriano/genética , ADN Bacteriano/metabolismo , ADN de Hongos/genética , ADN de Hongos/metabolismo , Escherichia coli/clasificación , Escherichia coli/efectos de los fármacos , Escherichia coli/genética , Escherichia coli/metabolismo , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Mutación , Saccharomyces cerevisiae/clasificación , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/metabolismo , Schizosaccharomyces/clasificación , Schizosaccharomyces/efectos de los fármacos , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Zymomonas/clasificación , Zymomonas/efectos de los fármacos , Zymomonas/genética , Zymomonas/metabolismo
17.
Sci Rep ; 8(1): 14629, 2018 10 02.
Artículo en Inglés | MEDLINE | ID: mdl-30279451

RESUMEN

The fission yeast clade, which has a distinct life history from other yeasts, can provide important clues about evolutionary changes. To reveal these changes the large S. cryophilus supercontigs were assembled into chromosomes using synteny relationships and the conserved pericentromeric, subtelomeric genes. Togetherness of the supercontigs was confirmed by PCR. Investigation of the gene order revealed localisation of the rDNA arrays, more than 300 new conserved orthologues and proved that S. cryophilus supercontigs were mosaics of collinear blocks. PFGE analysis showed that size of the S. cryophilus chromosomes differ from the S. pombe chromosomes. Comparative genomic analyses of the newly assembled chromosomes confirmed that the closest relative of S. cryophilus was S. octosporus not just in sequence similarity but also in a structural way, and revealed that preservation of the conserved regions did not arise from the lower number of chromosomal rearrangements. Translocations were more typical in the closely related species, while the number of inversions increased with the phylogenetic distances. Our data suggested that sites of the chromosomal rearrangements were not random and often associated with repetitive sequences, structural- and nucleotide evolution might correlate. Chromosomal rearrangements of the fission yeasts compared to other lineages were also discussed.


Asunto(s)
Evolución Molecular , Genoma Fúngico , Schizosaccharomyces/clasificación , Schizosaccharomyces/genética , Cromosomas Fúngicos , Orden Génico , Genómica/métodos , Haploidia , Filogenia
18.
PLoS One ; 11(3): e0151102, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27007548

RESUMEN

At present, wine is generally produced using Saccharomyces yeast followed by Oenococus bacteria to complete malolactic fermentation. This method has some unsolved problems, such as the management of highly acidic musts and the production of potentially toxic products including biogenic amines and ethyl carbamate. Here we explore the potential of the fission yeast Schizosaccharomyces pombe to solve these problems. We characterise an extensive worldwide collection of S. pombe strains according to classic biochemical parameters of oenological interest. We identify three genetically different S. pombe strains that appear suitable for winemaking. These strains compare favourably to standard Saccharomyces cerevisiae winemaking strains, in that they perform effective malic acid deacidification and significantly reduce levels of biogenic amines and ethyl carbamate precursors without the need for any secondary bacterial malolactic fermentation. These findings indicate that the use of certain S. pombe strains could be advantageous for winemaking in regions where malic acid is problematic, and these strains also show superior performance with respect to food safety.


Asunto(s)
Schizosaccharomyces/metabolismo , Vino , Fermentación , Schizosaccharomyces/clasificación , Schizosaccharomyces/genética , Especificidad de la Especie
19.
Cytogenet Genome Res ; 110(1-4): 566-74, 2005.
Artículo en Inglés | MEDLINE | ID: mdl-16093710

RESUMEN

Recent studies of the LTR-retrotransposons of Schizosaccharomyces pombe have shed considerable light on their evolution and function. The sequencing of the S. pombe genome allowed analysis of its transposon content. This analysis provides information about the maintenance and loss of transposons in the genome. The results of transposition assays and biochemical analyses demonstrate that the N-terminal protein of Tf1 is functionally equivalent to the Gag proteins of retroviruses and retrotransposons. Despite this conservation of function, the N-terminal protein of Tf1 lacks any sequence similarity to other known Gag proteins. Sequence analysis and experimental data also indicate that the Tf1 transposons of S. pombe target their integration into specific sites in the host genome. Transposition events resulting from the expression of Tf1 reveal a strong preference for intergenic regions, specifically at pol II promoters in a window 100-400 bp upstream of open reading frames. The complete and partial copies of Tf transposons in the sequenced genome of S. pombe show the same association of integration with promoter regions. This body of work explores how the transposon interacts with the host, the balance between the transposons propagation and loss, and how different families of transposons evolve.


Asunto(s)
ADN Polimerasa II/genética , Elementos Transponibles de ADN , Evolución Molecular , Schizosaccharomyces/genética , Intrones , Sistemas de Lectura Abierta , Filogenia , Regiones Promotoras Genéticas , Retroelementos , Schizosaccharomyces/clasificación , Schizosaccharomyces/enzimología , Secuencias Repetidas Terminales
20.
Artículo en Ruso | MEDLINE | ID: mdl-16028519

RESUMEN

The cultural, morphological, physiological and biological characteristics of Schizosaccharomyces hominis, Benedek strain BKM Y-650 were under study. This name is regarded as synonymous to Schizosaccharomyces pombe Lindner.


Asunto(s)
Schizosaccharomyces/clasificación , Schizosaccharomyces/fisiología , Animales , Humanos , Micosis/microbiología , Schizosaccharomyces/citología
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