RESUMEN
Plant pathogenic and food spoilage microorganisms cause serious losses in crop production and severe damage during food manufacturing, transportation and storage. Synthetic antimicrobial agents are commonly used to control their propagation and harmful activities. However, the recent trend is shifting from chemicals towards safer and more eco-friendly alternatives. The use of antagonistic microorganisms as biological antimicrobial agents is becoming popular throughout the world to replace chemical agents. High numbers of microorganisms have turned out to exert adverse/inhibitory effects on other microorganisms including pathogens and spoiling strains. However, most of them are only active under laboratory conditions and their activity is sensitive to environmental changes. Only a small number of them can be used to manufacture biological protective products on an industrial scale. Therefore, there is a great need to identify additional antagonists. Yeasts have come to the forefront of attention because antimicrobial antagonism is fairly widespread among them. In the recent years, numerous excellent review articles covered various aspects of the phenomenon of antimicrobial antagonism of yeasts. However, none of them dealt with how antagonistic yeasts can be sought and identified, despite the high number and diverse efficiency of screening and identification procedures. As researchers working in different laboratories use different criteria and different experimental set-ups, a yeast strain found antagonistic in one laboratory may prove to be non-antagonistic in another laboratory. This review aims to provide a comprehensive and partially critical overview of the wide diversity of identification criteria and procedures to help researchers choose appropriate screening and identification strategies.
RESUMEN
Saccharomyces strains with chimerical genomes consisting of mosaics of the genomes of different species ("natural hybrids") occur quite frequently among industrial and wine strains. The most widely endorsed hypothesis is that the mosaics are introgressions acquired via hybridisation and repeated backcrosses of the hybrids with one of the parental species. However, the interspecies hybrids are sterile, unable to mate with their parents. Here, we show by analysing synthetic Saccharomyces kudriavzevii x Saccharomyces uvarum hybrids that mosaic (chimeric) genomes can arise without introgressive backcrosses. These species are biologically separated by a double sterility barrier (sterility of allodiploids and F1 sterility of allotetraploids). F1 sterility is due to the diploidisation of the tetraploid meiosis resulting in MAT a /MAT α heterozygosity which suppresses mating in the spores. This barrier can occasionally be broken down by malsegregation of autosyndetically paired chromosomes carrying the MAT loci (loss of MAT heterozygosity). Subsequent malsegregation of additional autosyndetically paired chromosomes and occasional allosyndetic interactions chimerise the hybrid genome. Chromosomes are preferentially lost from the S. kudriavzevii subgenome. The uniparental transmission of the mitochondrial DNA to the hybrids indicates that nucleo-mitochondrial interactions might affect the direction of the genomic changes. We propose the name GARMe (Genome AutoReduction in Meiosis) for this process of genome reduction and chimerisation which involves no introgressive backcrossings. It opens a way to transfer genetic information between species and thus to get one step ahead after hybridisation in the production of yeast strains with beneficial combinations of properties of different species.
Asunto(s)
Genoma Fúngico , Hibridación Genética , Meiosis , Recombinación Genética , Saccharomyces/genética , ADN de Hongos/genética , ADN Mitocondrial , Genómica , Polimorfismo de Longitud del Fragmento de Restricción , Análisis de Secuencia de ADN , Vino/microbiologíaRESUMEN
Simple and efficient genotyping methods are widely used to assess the diversity of a large number of microbial strains, e.g. wine yeasts isolated from a specific geographical area or a vintage. Such methods are often also the first to be applied, to decrease the number of strains deemed interesting for a more time-consuming physiological characterization. Here, we aimed to use a physiologically characterized strain collection of 69 Saccharomyces cerevisiae strains from Hungarian wine regions to determine whether geographical origin or physiological similarity can be recovered by clustering the strains with one or two simultaneously used variations of interdelta genotyping. Our results indicate that although a detailed clustering with high resolution can be achieved with this method, the clustering of strains is largely contrasting when different primer sets are used and it does not recover geographical or physiological groups. SIGNIFICANCE AND IMPACT OF THE STUDY: Genotyping is routinely used for assessing the diversity of a large number of isolates/strains of a single species, e.g. a collection of wine yeasts. We tested the efficiency of interdelta genotyping on a collection of Saccharomyces wine yeasts from four wine regions of Hungary that was previously characterized physiologically. Interdelta fingerprinting recovered neither physiological nor geographical similarities, and in addition, the two different primer pairs widely used for this method showed conflicting and barely comparable results. Thus, this method does not necessarily represent the true diversity of a strain collection, but detailed clustering may be achieved by the combined use of primer sets.
Asunto(s)
Saccharomyces cerevisiae/aislamiento & purificación , Vino/microbiología , Biodiversidad , Fermentación , Genotipo , Hungría , Saccharomyces cerevisiae/clasificación , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Vino/análisisRESUMEN
Combination of molecular genetic analysis (karyotyping, PCR-RFLP of MET2, the ITS1-ITS2 region and the NTS region) and physiological examination (melibiose and mannitol utilization, sugar-, ethanol- and copper tolerance, killer activity, fermentation vigor and production of metabolites) of yeasts isolated from spontaneously fermenting wines in four wine regions revealed very high diversity in the Saccharomyces cerevisiae populations. Practically each S. cerevisiae isolate showed a unique pattern of properties. Although the strains originating from the same wine were quite similar in certain traits, they showed diversity in other properties. These results indicate that alcoholic fermentation in grape wines is performed by highly diverse yeast consortia rather than by one or two dominating strains. The less frequent Saccharomyces uvarum strains were less diverse, showed lower karyotype variability, were Mel(+), Man(+), more sensitive to 60% sugar, and ethanol or copper in the medium. They produced less acetic acid and fermented better at 14 degrees C than most of the S. cerevisiae isolates, but certain S. cerevisiae strains showed comparably high fermentation rates at this temperature, indicating that it is not a general rule that S. uvarum ferments better than S. cerevisiae at low temperatures. The segregation of certain traits (melibiose utilization, mannitol utilization and copper resistance) in both species indicates that the genomes can easily change during vegetative propagation. The higher diversity among the S. cerevisiae isolates suggests that the S. cerevisiae genome may be more flexible than the S. uvarum genome and may allow more efficient adaptation to the continuously changing environment in the fermenting wine.
Asunto(s)
Biodiversidad , Fermentación , Saccharomyces/genética , Saccharomyces/metabolismo , Vino/microbiología , Genotipo , Hungría , Datos de Secuencia Molecular , Fenotipo , Filogenia , Saccharomyces/clasificación , Saccharomyces/aislamiento & purificación , Vino/análisisRESUMEN
Minimal inhibitory and minimal fungicidal concentrations of caspofungin were determined for 48 Candida inconspicua isolates. By using CLSI (formerly NCCLS) methodology with the partial inhibition endpoint criterion, caspofungin exhibited a good fungicidal effect against C. inconspicua (the MIC(90) was 0.25 microg/ml and the minimum fungicidal concentration [MFC] was 0.5 microg/ml after 24 h). Total inhibition yielded falsely elevated MICs, exceeding even the respective MFCs.
Asunto(s)
Antifúngicos/farmacología , Candida/efectos de los fármacos , Péptidos Cíclicos/farmacología , Caspofungina , Relación Dosis-Respuesta a Droga , Farmacorresistencia Fúngica , Equinocandinas , Fluconazol/farmacología , Humanos , Lipopéptidos , Pruebas de Sensibilidad Microbiana/métodos , Pruebas de Sensibilidad Microbiana/normasRESUMEN
The fork-head type transcription factors are a class of regulators that function in a broad spectrum of cellular and developmental processes in many species ranging from yeasts to human. Previous data on yeast fork-head genes suggested roles for these regulators in the control of cell division, sexual differentiation and development. The genome of Schizosaccharomyces pombe has four genes that code for proteins containing fork-head domains (FKH), two of which have been characterised. Here we describe the remaining two genes, fhl1 and fkh2, that code for proteins containing fork-head-associated domains (FHA) besides their FKHs. Neither of them is essential for viability, although the deletion of either fhl1 (putative homologue of Saccharomyces cerevisiae FHL1) or fkh2 (similar to FKH1 and FKH2 of S. cerevisiae) reduced the growth rate and caused an extension of cell length due to delayed G2-to-M transition. Occasionally, multiseptate cells were also produced, indicating the involvement of fhl1 and fkh2 in efficient septum cleavage. The fkh2Delta cells were slightly more sensitive than the wild-type cells to certain environmental stresses, showed reduced fertility and occasional deficiencies in meiosis II, indicating that fkh2 might also act in stress response and sexual differentiation.
Asunto(s)
Proteínas Nucleares/genética , Proteínas de Schizosaccharomyces pombe/genética , Schizosaccharomyces/genética , Factores de Transcripción/genética , Secuencia de Aminoácidos , Compuestos de Cadmio/farmacología , Ciclo Celular/genética , División Celular/efectos de los fármacos , División Celular/genética , Relación Dosis-Respuesta a Droga , Factores de Transcripción Forkhead , Fase G2/genética , Genoma Fúngico , Calor , Peróxido de Hidrógeno/farmacología , Meiosis/genética , Metilmetanosulfonato/farmacología , Datos de Secuencia Molecular , Filogenia , Schizosaccharomyces/efectos de los fármacos , Schizosaccharomyces/fisiología , Alineación de Secuencia , Eliminación de Secuencia , Homología de Secuencia de Aminoácido , Sorbitol/farmacología , Esporas Fúngicas/genética , Sulfatos/farmacologíaRESUMEN
AIMS: Although numerous physiological and molecular methods have been proposed for yeast taxonomy, the unambiguous separation of Saccharomyces sensu stricto species in natural samples is still an incompletely resolved issue. In this study the power of various methods was compared in the identification of strains isolated from fermenting botrytized grape musts. METHODS AND RESULTS: Conventional taxonomic and physiological tests and molecular methods developed for rapid identification were used. CONCLUSIONS: None of the methods tested was sufficiently powerful. However, the combination of electrophoretic karyotyping and the PCR-RFLP of MET2 with growth tests at 10 and 37 degrees C provided results sufficient for species identification of Saccharomyces wine strains which were not interspecific hybrids or recombinants. SIGNIFICANCE AND IMPACT OF THE STUDY: The proposed combination of molecular and physiological methods allows specific taxonomic identification and separation of Saccharomyces wine strains without extensive genetic and molecular analysis. The proposed combined approach can also identify hybrids and recombinants.
Asunto(s)
Microbiología de Alimentos , Saccharomyces/clasificación , Vitis/microbiología , Vino/microbiología , Cromosomas Fúngicos/genética , Desoxirribonucleasas de Localización Especificada Tipo II/genética , Electroforesis/métodos , Fermentación , Cariotipificación/métodos , Reacción en Cadena de la Polimerasa/métodos , Polimorfismo de Longitud del Fragmento de Restricción , Ribotipificación/métodos , Saccharomyces/crecimiento & desarrollo , Saccharomyces/aislamiento & purificación , Saccharomyces cerevisiae/clasificación , Saccharomyces cerevisiae/crecimiento & desarrollo , Saccharomyces cerevisiae/aislamiento & purificación , TemperaturaRESUMEN
We present data demonstrating that the strain DBVPG 3827 does not belong to C. stellata. From the results of physiological analysis, electrophoretic karyotyping, the PCR-RFLP of the ITS1-5.8S-ITS2 region and the nucleotide sequence of the D1/D2 domain of the 26S rDNA it can be concluded that DBVPG 3827 is a strain of Starmerella bombicola. This finding, and the recent observation that C. stellata can easily be confused with C. zemplinina in tests of conventional taxonomy, urges a critical revision of the enological role(s) attributed by researchers to C. stellata.
Asunto(s)
Ascomicetos/clasificación , Candida/clasificación , Ascomicetos/genética , Ascomicetos/crecimiento & desarrollo , Ascomicetos/fisiología , Candida/genética , Candida/crecimiento & desarrollo , Candida/fisiología , ADN de Hongos/análisis , ADN Espaciador Ribosómico/análisis , Cariotipificación , Técnicas de Tipificación Micológica , Filogenia , Reacción en Cadena de la Polimerasa , Polimorfismo de Longitud del Fragmento de Restricción , ARN Ribosómico/genética , ARN Ribosómico 5.8S , Análisis de Secuencia de ADNRESUMEN
AIMS: Investigation of the meiotic segregation of karyotypes and physiological traits in indigenous Saccharomyces strains isolated from Aglianico (South Italy) red wine. METHODS AND RESULTS: Segregation was studied in F1 and F2 descendants. Tetrads were isolated from sporulating cultures by micromanipulation. The spore clones were subjected to karyotype analysis by pulse-field gel electrophoresis (Bio-Rad model CHEF-DR II) and to various physiological tests. Certain chromosomes of the isolates showed 2:2 segregation patterns in F1 but proved to be stable in F2. The ability of cells to utilize maltose also segregated in a 2 : 2 manner in F1 and did not segregate in F2. Resistance to CuSO4, SO2 tolerance, the fermentative power and the production of certain metabolites segregated in both F1 and F2 generations and showed patterns indicating the involvement of polygenic regulation. CONCLUSIONS: The analysis revealed a high degree of genetic instability and demonstrated that meiosis can improve chromosomal and genetic stability. SIGNIFICANCE AND IMPACT OF THE STUDY: Winemaking is critically dependent on the physiological properties and genetic stability of the fermenting Saccharomyces yeasts. Selection of clones from F2 or later generations can be a method of reduction of genetic instability.
Asunto(s)
Microbiología de Alimentos , Saccharomyces cerevisiae/fisiología , Vino/microbiología , Cromosomas Fúngicos/genética , Cobre/metabolismo , Medios de Cultivo , Etanol/metabolismo , Fermentación/genética , Galactosa/metabolismo , Cariotipificación/métodos , Maltosa/metabolismo , Meiosis/genética , Análisis de Componente Principal/métodos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismoRESUMEN
Stable chromium(VI)-sensitive and -tolerant mutants were obtained by induced mutagenesis of Schizosaccharomyces pombe lysine and leucine auxotrophic heterothallic strains 6chr+ and 9chr+. Eleven of them were selected for further studies. Fast transport of 51CrO4(2-) was detected in a representative sensitive mutant, chr-51S, while the tolerant mutant chr1-66T and the parental strain 6chr+ exhibited significantly lower 51CrO4(2-) uptake. The segregation of tetrads of three selected CrVI-tolerant mutants, chr1-66T, chr1-14T and chr2-04T, strongly indicated that tolerance was determined by single mutations. Random spore analysis proved that the mutations of chr1-66T and chr1-14T were allelic and the mutation of mutant chr2-04T was not allelic with the mutation of chr1-66T. Recombinants carrying the ura4D18 selective marker were created for transformation experiments. Two of them (chr1-661T and chr2-046T) can be used to clone and identify the genes responsible for their CrVI tolerance phenotype.
Asunto(s)
Cromo/metabolismo , Cromo/farmacología , Farmacorresistencia Fúngica/genética , Schizosaccharomyces/genética , Schizosaccharomyces/metabolismo , Alelos , Antifúngicos/metabolismo , Antifúngicos/farmacología , Transporte Biológico , Recuento de Colonia Microbiana , Cruzamientos Genéticos , Genes Dominantes , Genes Fúngicos , Prueba de Complementación Genética , Cinética , Metales/metabolismo , Pruebas de Sensibilidad Microbiana , Mutación , Recombinación Genética , Schizosaccharomyces/efectos de los fármacos , Schizosaccharomyces/crecimiento & desarrollo , Transformación GenéticaRESUMEN
We previously identified four nuclear genes (caf1+-caf4+) in Schizosaccharomyces pombe, mutations in which confer resistance to caffeine and brefeldin A. caf1+, caf2+ and caf4+ were sequenced and found to be identical to the multidrug-resistance/stress-response genes hba1, crm1 and trr1, respectively. Here we show that caf3 is allelic to pap1, which encodes an AP-1-like transcription factor. The allele associated with caffeine resistance, caf3-89, contains a single-nucleotide exchange that results in a Leu-->Ser exchange in the NES (nuclear export signal) domain of the gene product. Due to this alteration, the modified protein can not be exported from the nucleus back into the cytoplasm, and thus accumulates in the nucleus. The activity of pap1/caf3 is shown to be necessary for manifestation of the caffeine resistance caused by mutations in the genes hba1/caf1 and crm1/caf2. We also cloned two genes that confer caffeine resistance when carried on a multicopy plasmid. One of them turned out to be a truncated allele of pad1/bfr2/sks1, which codes for a subunit of the 26 S proteosome. The putative product of the other gene, designated caf5, has a structure highly similar to that of MFS permeases. It contains two groups of six transmembrane spanning domains each, with the conserved motifs WRW, PET and GAIGGPVLGP in the fifth and sixth domains. These results are all consistent with our earlier hypothesis, which suggested that the caf genes are functionally interlinked in a complex detoxification mechanism. caf5 and pad1 may also encode parts of this mechanism.
Asunto(s)
Cafeína , Proteínas de Unión al ADN/genética , Farmacorresistencia Fúngica Múltiple/genética , Proteínas Fúngicas/genética , Proteínas de la Membrana/genética , Complejo de la Endopetidasa Proteasomal , Proteínas de Schizosaccharomyces pombe/genética , Schizosaccharomyces/genética , Factores de Transcripción/genética , Alelos , Secuencia de Aminoácidos , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico , Peróxido de Hidrógeno , Datos de Secuencia Molecular , Mutación Missense/genética , Proteínas Asociadas a Pancreatitis , Péptido Hidrolasas/genética , Filogenia , Plásmidos/genética , Alineación de Secuencia , Análisis de Secuencia de ADN , Factores de Transcripción/fisiologíaRESUMEN
We have previously described the genetic analysis of eleven complementation groups ( sep6- sep16) defined by Schizosaccharomyces pombe mutants that are defective in cell separation and sexual differentiation. Here we report on the cloning and characterisation of two members of this set, sep10 and sep11. Sequencing of the full-length sep10 revealed a continuous ORF that encodes a conserved protein with possible functions in general transcriptional regulation. The coding region of sep11 is interrupted by introns and the putative s ep11 protein shows no sequence similarity with known proteins of other species. Disruption of each gene causes temperature sensitivity. Simultaneous disruption of both genes is lethal, demonstrating that sep10 and sep11 perform related, overlapping functions. Overexpression of aff1/ste11, a pivotal regulator of sexual development, suppresses the sterility of sep10 (-) cells, which suggests that sep10 is needed for the activity of aff1/ste11.
Asunto(s)
Genes Fúngicos/fisiología , Proteínas de Schizosaccharomyces pombe/genética , Schizosaccharomyces/genética , Factores de Transcripción/genética , Secuencia de Aminoácidos , Clonación Molecular , Reparación del ADN , ADN de Hongos , Regulación Fúngica de la Expresión Génica , Genes Letales , Mitosis/genética , Datos de Secuencia Molecular , Mutación , Sistemas de Lectura Abierta , Fenotipo , Recombinación Genética , Proteínas de Schizosaccharomyces pombe/metabolismo , Homología de Secuencia de AminoácidoRESUMEN
Using genetic hybridisation analysis and molecular karyotyping we revealed an association of Saccharomyces bayanus var. uvarum species with Tokaj wine-making. Along with identification of Saccharomyces strains isolated by E. Minárik in Slovakia, the composition of Tokaj populations in Hungary was studied. Twenty-eight Hungarian Saccharomyces strains were analysed in terms of karyotype. The majority of strains belong to S. bayanus var. uvarum. Two non-identified Saccharomyces strains were found to be polyploid according to their complex karyotype patterns.
Asunto(s)
Saccharomyces/clasificación , Vino/microbiología , ADN Bacteriano/química , ADN Bacteriano/genética , Electroforesis en Gel de Campo Pulsado , Hungría , Cariotipificación , Saccharomyces/genética , Saccharomyces/metabolismo , EslovaquiaAsunto(s)
Proteínas de Ciclo Celular/genética , Regulación Fúngica de la Expresión Génica , Proteínas de Schizosaccharomyces pombe/genética , Schizosaccharomyces/citología , Transcripción Genética , Proteínas de Ciclo Celular/metabolismo , División Celular , Schizosaccharomyces/genética , Proteínas de Schizosaccharomyces pombe/metabolismoRESUMEN
The dimorphic fission yeast Schizosaccharomyces japonicus has proved to be an excellent experimental model for the investigation of the eukaryotic cell. Here we show that it has a haplontic life cycle, in which the diploid phase is confined to the zygote. To make it amenable to genetic and molecular analysis, we generated genetic markers and cloned a genomic sequence which acts as ars when integrated into a plasmid. Diploids suitable for testing complementation and recombination between markers can be formed by protoplast fusion. The complementation tests and the recombination frequencies determined in octads of spores identified 28 non-allelic groups (genes) of mutations of the auxotrophic and mycelium-negative mutants. Two groups of linked markers were also identified. The cloned fragment, which expresses ars activity, encodes a putative amino acid sequence highly similar to a conserved domain of proteins Cut1 (Schizosaccharomyces pombe), BimB (Aspergillus nidulans) and Esp1 (Saccharomyces cerevisiae).
Asunto(s)
Proteínas de Ciclo Celular/genética , Replicación del ADN , ADN de Hongos , Proteínas de Schizosaccharomyces pombe , Schizosaccharomyces/genética , Secuencia de Aminoácidos , Secuencia de Bases , Clonación Molecular , Proteínas Fúngicas/genética , Genes Fúngicos , Marcadores Genéticos , Datos de Secuencia Molecular , Mutación , Micelio/metabolismo , Schizosaccharomyces/crecimiento & desarrollo , Schizosaccharomyces/metabolismo , Alineación de Secuencia , Esporas Fúngicas/metabolismoRESUMEN
The proper division of cells is essential for the production of viable daughter cells. In plants and fungi, the dividing cell produces a cross-wall or septum that bisects the cytoplasm. For separation of the daughter cells, the septum has to be cleaved. To study the regulation of this process, we isolated mutants defective in septum cleavage. The mutants showed highly pleiotropic phenotypes and defined 17 novel genes. The deduced amino acid sequences of the products of the cloned genes exhibited homologies to various transcription regulators of other organisms. The homologies and the pleiotropic effects of the mutations on sexual development, stress response, mitotic stability, septum initiation and septum placement indicated that these genes affect cell separation indirectly, through multifunctional regulatory modules.
Asunto(s)
Genes Fúngicos , Schizosaccharomyces/citología , Schizosaccharomyces/genética , División CelularRESUMEN
rec7 is involved in intra- and intergenic meiotic recombination in all tested regions of the genome of the fission yeast Schizosaccharomyces pombe. Segregational analysis in a rec7 gene disruption mutant revealed frequent occurrence of two-spored asci. Spores giving rise to diploid colonies were shown to derive from skipping of the second meiotic division. Nondisjunction of homologous chromosomes at the first meiotic division was also frequent. The cytological structures and processes, such as formation of linear elements, pairing of homologous chromosomes, and clustering of telomeres and centromeres, are regular in the mutant. Northern blot experiments revealed meiosis-specific expression of rec7. Screening of a meiotic cDNA library also identified transcripts from the opposite strand in the rec7 region. A Rec7-GFP fusion protein was localized in the nucleus of whole cells before karyogamy, during prophase, and after meiosis I. On spreads of prophase nuclei approximately 50 foci of Rec7-GFP were counted. Some of the observed phenotypes of the disruption mutant and the N-terminal sequence homology suggest that Rec7p is a functional homolog of Rec114p of Saccharomyces cerevisiae. The observed phenotypes of the disruption and the appearance of Rec7-GFP in mating haploid cells and after meiosis I are consistent with Rec7p functions before, during, and after meiotic prophase.
Asunto(s)
Proteínas Fúngicas/genética , Meiosis , Recombinación Genética , Proteínas de Schizosaccharomyces pombe , Schizosaccharomyces/genética , Northern Blotting , División Celular , Núcleo Celular , Segregación Cromosómica , ADN Complementario/metabolismo , Proteínas Fúngicas/biosíntesis , Proteínas Fúngicas/química , Proteínas Fúngicas/fisiología , Biblioteca de Genes , Prueba de Complementación Genética , Genotipo , Proteínas Fluorescentes Verdes , Homocigoto , Proteínas Luminiscentes/metabolismo , Modelos Genéticos , Mutagénesis , No Disyunción Genética , Fenotipo , Proteínas Recombinantes de Fusión/metabolismo , Factores de TiempoRESUMEN
The mediator complexes transduce regulatory information from upstream regulatory elements to the transcription machinery in organisms ranging from yeasts to humans. By a genome-wide search we identified 14 ORFs and genes in the genome of the fission yeast Schizosaccharomyces pombe that encode putative homologues of Saccharomyces cerevisiae mediator subunits. The Sch. pombe proteins are smaller and appear to form a mediator of lower complexity, which is consistent with the hypothesized ancient origin of fission yeasts.
Asunto(s)
Proteínas Fúngicas/genética , Genes Fúngicos , Saccharomyces cerevisiae/genética , Schizosaccharomyces/genética , Factores de Transcripción/genética , Secuencia de Aminoácidos , Proteínas Fúngicas/química , Genoma Fúngico , Humanos , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Homología de Secuencia de Aminoácido , Especificidad de la Especie , Factores de Transcripción/químicaRESUMEN
The Schizosaccharomyces pombe sep1 gene encodes a putative transcription factor that is required for cell separation. Among the genes required for septum formation and cytokinesis in fission yeast examined to date, the only one whose mRNA fluctuates significantly during the cell cycle is cdc15. In this study we have examined cdc15 mRNA levels in sep1 mutant and null backgrounds and have found that sep1p function is required for periodic accumulation of cdc15 mRNA. We have also localised sep1p and find that it is a nuclear protein, consistent with its proposed role as a transcription factor.
Asunto(s)
Proteínas de Ciclo Celular/genética , Exorribonucleasas/metabolismo , Proteínas de Unión al GTP/genética , Regulación Fúngica de la Expresión Génica , Proteínas Nucleares/metabolismo , Periodicidad , Proteínas de Saccharomyces cerevisiae , Schizosaccharomyces/genética , Ciclo Celular , Núcleo Celular/química , Clonación Molecular , Exorribonucleasas/genética , Eliminación de Gen , Genes Esenciales , Proteínas Nucleares/genética , ARN de Hongos/genética , ARN de Hongos/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Schizosaccharomyces/citología , Schizosaccharomyces/enzimologíaRESUMEN
The establishment of growth polarity in Schizosaccharomyces pombe cells is a combined function of the cytoplasmic cytoskeleton and the shape of the cell wall inherited from the mother cell. The septum that divides the cylindrical cell into two siblings is formed midway between the growing poles and perpendicularly to the axis that connects them. Since the daughter cells also extend at their ends and form their septa at right angles to the longitudinal axis, their septal (division) planes lie parallel to those of the mother cell. To gain a better understanding of how this regularity is ensured, we investigated septation in spherical cells that do not inherit morphologically predetermined cell ends to establish poles for growth. We studied four mutants (defining four novel genes), over 95% of whose cells displayed a completely spherical morphology and a deficiency in mating and showed a random distribution of cytoplasmic microtubules, Tea1p, and F-actin, indicating that the cytoplasmic cytoskeleton was poorly polarized or apolar. Septum positioning was examined by visualizing septa and division scars by calcofluor staining and by the analysis of electron microscopic images. Freeze-substitution, freeze-etching, and scanning electron microscopy were used. We found that the elongated bipolar shape is not essential for the determination of a division plane that can separate the postmitotic nuclei. However, it seems to be necessary for the maintenance of the parallel orientation of septa over the generations. In the spherical cells, the division scars and septa usually lie at angles to each other on the cell surface. We hypothesize that the shape of the cell indirectly affects the positioning of the septum by directing the extension of the spindle.