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M Sipiczki

Publications and source records attributed to M Sipiczki.

At least 19 recordsLinked to original sources
PubMed2005

Characterisation of two novel fork-head gene homologues of Schizosaccharomyces pombe: their involvement in cell cycle and sexual differentiation.

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.

Amino Acid Sequence↗
PubMed2005

Taxonomic reclassification of Candida stellata DBVPG 3827.

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.

Ascomycota↗
PubMed2005

Combined application of methods to taxonomic identification of Saccharomyces strains in fermenting botrytized grape must.

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.

Chromosomes, Fungal↗
PubMed2005

Caspofungin susceptibility testing of Candida inconspicua: correlation of different methods with the minimal fungicidal concentration.

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.

Antifungal Agents↗
PubMed2004

The transcription factor Pap1/Caf3 plays a central role in the determination of caffeine resistance in Schizosaccharomyces pombe.

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.

Alleles↗
PubMed2004

Characterization of chromate-sensitive and -tolerant mutants of Schizosaccharomyces pombe.

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.

Alleles↗
PubMed2004

Genetic segregation of natural Saccharomyces cerevisiae strains derived from spontaneous fermentation of Aglianico wine.

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.

Chromosomes, Fungal↗
PubMed2002

The Schizosaccharomyces pombe genes sep10 and sep11 encode putative general transcriptional regulators involved in multiple cellular processes.

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.

Amino Acid Sequence↗
PubMed2002

Saccharomyces bayanus var. uvarum in Tokaj wine-making of Slovakia and Hungary.

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.

DNA, Bacterial↗
PubMed2002

Marker construction and cloning of a cut1-like sequence with ARS activity in the fission yeast Schizosaccharomyces japonicus.

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).

Amino Acid Sequence↗
PubMed2001

Characterization of rec7, an early meiotic recombination gene in Schizosaccharomyces pombe.

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.

Blotting, Northern↗
PubMed2001

Multifunctional cytokinesis genes in Schizosaccharomyces pombe.

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.

Cell Division↗
PubMed2001

Identification of Schizosaccharomyces pombe genes that encode putative homologues of Saccharomyces cerevisiae mediator complex subunits.

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.

Amino Acid Sequence↗
PubMed2000

The S. pombe sep1 gene encodes a nuclear protein that is required for periodic expression of the cdc15 gene.

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.

Cell Cycle↗
PubMed2000

Where does fission yeast sit on the tree of life?

The budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe are as different from each other as either is from animals: their ancestors separated about 420 to 330 million years ago. Now that S. pombe is poised to join the post-genome era, its evolutionary position should become much clearer.

Classification↗
PubMed2000

The use of morphomutants to investigate septum formation and cell separation in Schizosaccharomyces pombe.

Cytokinesis in the fission yeast Schizosaccharomyces begins by formation of a medially placed actomyosin ring, continues by progressive development of a septum, and is completed by cleavage of the mature septum to bring about cell separation. The cytological analysis of the Schizosaccharomyces pombe morphomutants sph2-3 and sep1-1 presented here demonstrates that the medial actomyosin ring colocalizes to the leading edge of the centripetally growing septum, and cleavage of the septum is triggered by rupture of the mother cell wall at the septal basis.

Actins↗
PubMed2000

The Schizosaccharomyces pombe sep15+ gene encodes a protein homologous to the Med8 subunit of the Saccharomyces cerevisiae transcriptional mediator complex.

We previously described the isolation of mutants defective in cell separation and the identification of 16 sep genes with complex functions. Here we report on the cloning and analysis of sep15+. The deduced amino acid sequence of the Sep15 protein shows significant homology to Med8, a component of the Saccharomyces cerevisiae transcription mediator complex. The mutation sep15-598 confers hyphal morphology and causes temperature-sensitive lethality. Disruption of sep15+ is lethal, indicating that Sep15 exerts an essential function and its role in cell separation is indirect.

Amino Acid Sequence↗