bluuebunny/crossref_metadata_2025 huggingface dataset:
Saccharomyces cerevisiae mata mutant cells defective in pointed projection formation in response to α‐factor at high concentrations
Authors: Yorihuzi Tetuya , Ohsumi Yoshinori
Published: May 2005

We have isolated Saccharomyces cerevisiae MATa mutant cells that do not form a pointed projection but elongate in response to α‐factor at high concentrations. Complementation tests defined three genes, PPF1, PPF2, and PPF3 (for pointed projection formation), necessary for pointed projection formation. Allelism tests with genes known to be needed for projection formation revealed that PPF1 is identical to SPA2, while PPF2 and PPF3 are not allelic to SST2, STE2, SPA2, BEM1 or SLK1/SSP31/BCK1. The morphology of MATa ppf mutants treated with high concentrations of α‐factor is similar to that of MATa PPF cells treated with α‐factor at low concentrations. Quantitative mating tests showed that PPF2 and PPF3 are not essential for mating in either MATa or MATα background. Monitoring of division arrest and expression of an α‐factor‐inducible gene revealed that mutations in the PPF genes do not affect the responses of MATa cells to low concentrations of α‐factor. Unlike wild‐type cells, the ppf mutants exhibited early recovery from α‐factor‐induced division arrest. Furthermore, vegetatively growing ppf3‐1 cells are slightly defective in cell separation of mother and daughter cells and in selection of the correct bud sites in all cell types. These results indicate that PPF2 and PPF3 are involved in the response to α‐factor at high concentrations and that PPF3 is also required for proper establishment of polarity in vegetative growth.
V. Yeast mapping reports. The MAG1 3‐methladenine DNA glycosylase gene is closely linked to the SPT15 TATA‐binding TFIID gene on chromosome V‐R in Saccharomyces cerevisiae
Authors: Xiao Wei , Penugonde Vasu , Rank Gerald H.
Published: May 2005

The MAG1 gene encodes a 3‐methyladenine DNA glycosylase, which is involved in DNA alkylation repair in Saccharomyces cerevisiae. The mag1 mutant is deficient in 3‐methyladenine DNA glycosylase activity and shows enhanced sensitivity to several monofunctional alkylating agents. MAG1 is allelic to MMS5. This gene has been previously located on chromosome V by chromosomal hybridization. We present physical and genetic mapping data here showing that the MAG1 gene is located on chromosome V‐R, proximal to and about 10 kilobase pairs away from the SPT15 gene coding for the yeast TATA‐binding protein TFIID.
Yeast exo‐β‐glucanases can be used as efficient and readily detectable reporter genes in Saccharomyces cerevisiae
Authors: Cid V. J. , Alvarez A. M. , Santos A. I. , Nombela C. , Sanchez M.
Published: May 2005

Yeast exo‐1,3‐β‐glucanases are secretable proteins whose function is basically trophic and may also be involved in cell wall glucan hydrolytic processes. Since fluorescein di(β‐D‐glucopyranoside) is a fluorogenic substrate detectable and quantifiable by flow cytometry, it was used for testing the ability of the EXG1 gene product of Saccharomyces cerevisiae and its homologous gene in Candida albicans to function as reporter genes. These open reading frames were coupled to different promoters in multicopy plasmids, and exoglucanase activity quantified at flow cytometry. Exoglucanases were found to be useful tools for the study of promoter regions in S. cerevisiae. This technique has the advantage over other reporter gene systems—such as β‐galactosidase fusions—that it does not require permeabilization of yeast cells and therefore it allows the recovery of viable cells—by sorting—after flow cytometry analysis.
Yeast sequencing reports. Comparison of INO1 gene sequences and products in Candida albicans and Saccharomyces cerevisiae
Authors: Klig Lisa S. , Zobel Pamela A. , Devry Christopher G. , Losberger Christoph
Published: May 2005

The sequence of the Candida albicans inositol biosynthetic gene, CaINO1, and its flanking regions is determined in this study. The largest open reading frame has a coding sequence of 1560 base pairs, corresponding to a predicted protein of 521 amino acids. Three primary transcriptional start sites are found 64, 57 and 52 base pairs upstream of the ATG translational start site at position 1374. Five stop codons exist in a cluster at the end of the coding region. Within the upstream region TATA and CAAT eukaryotic regulatory sequences are identified along with regions corresponding to a 10 base pair inositol/choline responsive element consensus sequence. Computer analysis of the DNA sequence shows strong homology to the Saccharomyces cerevisiae INO1 gene. A comparison of the deduced amino acid sequence of the C. albicans INO1 gene product, inositol‐1‐phosphate synthase, with its homolog in S. cerevisiae shows 64% identity and 77% similarity. The differences between the two proteins are most prominent in the N‐terminal regions. The sequence has been deposited in the GenBank/EMBL data library under Accession Number L22737.
XV. Yeast sequencing reports. Isolation and DNA sequence of the STE13 gene encoding dipeptidyl aminopeptidase
Authors: Anna‐arriola Sonia Santa , Herskowitz Ira
Published: May 2005

We have isolated a mutant which exhibits partial constitutivity for a‐specific gene expression in α cells. The wild‐type gene was cloned and demonstrated to be allelic to the STE13 gene, which encodes the dipeptidyl aminopeptidase involved in processing of the α‐factor prepropheromone. Thus, the mating defect of the ste13 mutations in α cells may result both from the production of incompletely processed α‐factor and from the increased expression of a‐specific genes.

The STE13 open reading frame of 931 amino acids contains a putative membrane‐spanning segment near its amino terminus and is 31% identical to a second yeast dipeptidyl aminopeptidase (DAP2). A null mutant of STE13 has been constructed: it is viable and sporulation‐proficient. The sequence has been deposited in the GenBank data library under Accession Number L21944.
Cloning and sequencing of Schizosaccharomyces pombe car1 gene encoding arginase. Expression of the arginine anabolic and catabolic genes in response to arginine and related metabolites
Authors: Van Huffel C. , Dubois E. , Messenguy F.
Published: May 2005

We report here the cloning and sequencing of the gene encoding arginase (car1) from Schizosaccharomyces pombe. Since no arginase‐less strain exists in this organism, we cloned the gene by functional complementation of a car1 mutant strain from Saccharomyces cerevisiae. The S. pombe car1 gene encodes a 323 amino acids polypeptide sharing identity with arginases from different organisms. Measurements of arg3, arg11 and car1 mRNA under different growth conditions confirm the very weak repression by arginine of the two anabolic genes and show that the induction of arginase synthesis operates at a transcriptional level. The promoter of S. pombe car1 gene does not contain the ‘arginine boxes’ defined as the target of the ARGR‐MCM1 proteins in the promoters of the arginine co‐regulated genes in S. cerevisiae. The heterologous expression of S. pombe car1 gene in S. cerevisiae is independent of the ARGRII gene product (ArgRIIp/Arg81p). Determination of arginine, ornithine and citrulline intracellular concentrations shows the efficiency of the different controls operating in S. cerevisiae, and also indicates that in S. pombe enzyme compartmentation is not always sufficient to control the arginine metabolic flux.
Activation of trehalase during growth induction by nitrogen sources in the yeast Saccharomyces cerevisiae depends on the free catalytic subunits of camp‐dependent protein kinase, but not on functional ras proteins
Authors: Durnez Peter , Pernambuco Maria Beatriz , Oris Els , Argüelles Juan‐Carlos , Mergelsberg Hubert , Thevelein Johan M.
Published: May 2005

Addition of a nitrogen‐source to glucose‐repressed, nitrogen‐starved G0 cells of the yeast Saccharomyces cerevisiae in the presence of a fermentable carbon source induces growth and causes within a few minutes a five‐fold, protein‐synthesis‐independent increase in the activity of trehalase. Nitrogen‐activated trehalase could be deactivated in vitro by alkaline phosphatase treatment, supporting the idea that the activation is triggered by phosphorylation. Yeast strains containing only one of the three TPK genes (which encode the catalytic subunit of cAMP‐dependent protein kinase) showed different degrees of nitrogen‐induced trehalase activation. The order of effectiveness was different from that previously reported for glucose‐induced activation of trehalase in glucose‐derepressed yeast cells. Further reduction of TPK‐encoded catalytic subunit activity by partially inactivating point mutations in the remaining TPK gene further diminished nitrogen‐induced trehalase activation, while deletion of the BCY1 gene (which encodes the regulatory subunit) in the same strains resulted in an increase in the extent of activation. Deletion of the RAS genes in such a tpkw1 bcy1 strain had no effect. These results are consistent with mediation of nitrogen‐induced trehalase activation by the free catalytic subunits alone. They support our previous conclusion that cAMP does not act as second messenger in this nitrogen‐induced activation process and our suggestion that a novel nitrogen‐induced signaling pathway integrates with the cAMP pathway at the level of the free catalytic subunits of protein kinase A. Western blot experiments showed that the differences in the extent of trehalase activation were not due to differences in trehalase expression. On the other hand, we cannot completely exclude that protein kinase A influences the nitrogen‐induced activation mechanism itself rather than acting directly on trehalase. However, any such alternative explanation requires the existence of an additional, yet unknown, mechanism for activation of trehalase besides the well‐established regulation by protein kinase A.
Analysis of β‐glucans and chitin in a Saccharomyces cerevisiae cell wall mutant using high‐performance liquid chromatography
Authors: Hong Zhi , Mann Paul , Shaw Karen J. , Didomenico Beth
Published: May 2005

We have previously shown that mutations in the yeast KNR4 gene resulted in pleiotropic cell wall defects, including resistance to killer 9 toxin, elevated osmotic sensitivity to SDS and increased resistance to zymolyase, a (1→3)‐β‐glucanase. In this report, we further demonstrated that knr4 mutant cells were more permeable to a chromogenic substrate, X‐GAL, suggesting that the mutant cell walls were leakier to certain non‐permeable molecules. To determine if these defects resulted from structural changes in the cell walls, we analysed the alkali‐insoluble cell wall components using HPLC assays developed for this purpose. Comparative analysis using four isogenic strains from a ‘knr4 disrupted’ tetrad demonstrated that mutant cell walls contained much less (1→3)‐β‐glucan and (1→6)‐β‐glucan; however, the level of chitin, a minor cell wall component, was found to be five times higher in the mutant strains compared to the wild‐type strains. The data suggested that the knr4 mutant cell walls were dramatically weakened, which may explain the pleiotropic cell wall defects.
VII. Yeast sequencing reports. Cloning and characterization of a sulphite‐resistance gene of Saccharomyces cerevisiae
Authors: Casalone Enrico , Colella Carlo M. , Daly Simona , Fontana Silvia , Torricelli Ilaria , Polsinelli Mario
Published: May 2005

In this paper we describe the cloning and sequencing of the gene (SUL1) responsible for sulphite resistance in a Saccharomyces cerevisiae mutant (Casalone et al., 1992). The deduced amino acid sequence predicted that the gene codes for a zinc‐finger protein with five fingers. Comparison of wild‐type and mutant gene sequences demonstrated that the mutation event was a transversion from C to G; as a consequence of the mutation a histidine substituted an aspartic acid, affecting directly the fourth finger structure. The SUL1 gene sequence corresponds to that of FZF1 gene (Breitwieser et al., 1993) to which no function was attributed. The sequence has been entered in the EMBL data library under Accession Number 77592.
Yeast sequencing reports. Cloning and DNA sequence of a Kluyveromyces lactis ERD1 homologue
Authors: Dean Neta
Published: May 2005

The ERD1 gene product is required for the correct localization of soluble proteins that normally reside in the endoplasmic reticulum (ER). Cell lacking ERD1 secrete resident ER proteins and, in addition, exhibit defects in the processing of glycoproteins. Here, the molecular characterization of the Kluyveromyces lactis ERD1 homologue is described. A comparison of the predicted sequences of the Saccharomyces cerevisiae and K. lactis Erd1 proteins indicates that they are about 30% identical and 50% similar in sequence. Despite low sequence identity, these proteins are predicted to be conserved structurally. Furthermore, the K. lactis protein can functionally complement an S. cerevisiae mutant containing a deletion of the entire ERD1 gene, indicating these two proteins are functional homologues. The GenBank data library Accession Number for the DNA sequence reported in this paper is UO4714.
A family of vectors that facilitate transposon and insertional mutagenesis of cloned genes in yeast
Authors: Allen James B. , Elledge Stephen J.
Published: May 2005

This report describes two sets of plasmid vectors that facilitate the identification of regions of complementation in cloned genomic inserts via transposon or insertional mutagenesis. The first set contains ARS‐H4 CEN6, a yeast selectable nutritional marker (HIS3, TRP1 or URA3), and neo for selection in Escherichia coli. These plasmids lack the Tn3 transposition immunity region present in pBR322 derived vectors, and are permissive recipients for Tn3 transposon mutagenesis. The second family of plasmids described facilitate gene disruption procedures performed in vitro. These vectors carry disruption cassettes that contain different yeast selectable markers (HIS3, LEU2, TRP1 or URA3) adjacent to the Tn5 neo gene. These genes can be excised as a cassette on a common restriction fragment and introduced into any desired restriction site with selection for kanamycin resistance.
Genetic and molecular analysis of hybrids in the genus Saccharomyces involving S. cerevisiae, S. uvarum and a new species, S. douglasii
Authors: Hawthorne Don , Philippsen Peter
Published: May 2005

We have studied the phenomenon of infertility of yeast hybrids obtained with physiological conditions under the control of compatible mating systems. The yeasts investigated are three Saccharomyces species: S. cerevisiae, S. uvarum and a new species, S. douglasii. The diploid hybrids from crosses between these species sporulate well but are essentially infertile. The rare viable spores, one per 104 to 105 asci, that have been examined carry a complete genome comprised of chromosomes contributed by both parents but invariably have extra chromosomes, i.e. they are generally disomic for at least two or three chromosomes. This observation is consistent with a failure, in meiosis I, of the pairing and disjunction of homologous chromosomes which in most cases results in spores with an incomplete set of chromosomes. This apparent lack of pairing of ‘homeologous’ chromosomes in meiosis I was analysed in most detail with S. cerevisiae/S. douglasii hybrids. As a genetic tool we studied frequencies of recombination, taking advantage of an S. douglasii breeding stock of some 50 identified mutations in non‐switching haploids. Recombination, although markedly reduced, could be observed at both the chromosomal and allelic levels, implying a sporadic pairing in meiosis to allow genetic exchange. Meiotic recombination frequencies were studied for 14 gene pairs and generally found to be reduced ten‐fold. Heteroallelic recombination (gene conversion) frequencies were measured at 22 loci and were judged to be reduced at least two‐ to 100‐fold. DNA hybridization experiments with S. cerevisiae gene probes gave results consistent with low DNA sequence homologies between S. cerevisiae and S. douglasii. Moreover, by chance, our experiments disclosed another Saccharomyces strain (CBS2908, originally classified as S. cerevisiae) with hybridization patterns identical to S. douglasii except for the hybridization with the Ty transposon probes. Crosses between CBS2908 and S. douglasii yielded diploid hybrids with 80–90% spore viability, thus establishing a second member of the S. douglasii species.
II. Yeast sequencing reports. Analysis of a 70kb region on the right arm of yeast chromosome II
Authors: Mannhaupt Gertrud , Stucka Rolf , Ehnle Susanne , Vetter Irene , Feldmann Horst
Published: May 2005

In the framework of the EC programme for sequencing yeast chromosome II, we have determined the nucleotide sequence of a 70 kb region. Subsequent analysis revealed 35 open reading frames, 14 of which correspond to known yeast genes. From structural parameters and/or similarity searches with entries in the current data libraries, a preliminary functional assessment of several of the putative novel gene products can be made. The gene density in this region amounts to one gene in 1.98 kb. Coding regions occupy 75% of the total DNA sequence. Within the intergenic regions, potential regulatory elements can be predicted. The data obtained here may serve as a basis for a more detailed biochemical analysis of the novel genes. The complete nucleotide sequence of the 70 kb segment as depicted in Figure 1 has been deposited in the EMBL data library under Accession Number X78993.
Efficient expression and secretion of recombinant alpha amylase in Pichia pastoris using two different signal sequences
Authors: Paifer E. , Margolles E. , Cremata J. , Montesino R. , Herrera L. , Delgado J. M.
Published: May 2005

We have cloned and expressed a bacterial thermostable alpha amylase gene in Pichia pastoris using the methanol‐controlled alcohol oxidase (AOX1) promoter. Two integrative vectors were constructed with two different secretion signal sequences in order to obtain efficient secretion of the protein. One vector contains the structural gene encoding the mature alpha amylase fused to the SUC2 gene signal sequence from Saccharomyces cerevisiae. In the other vector, the alpha amylase is expressed with its own signal sequence. In both cases, the alpha amylase were secreted into the culture medium with high efficiency, around 2·5 and 0·9 g/l respectively.
Characterization of lipid particles of the yeast, Saccharomyces cerevisiae
Authors: Leber Regina , Zinser Erwin , Paltauf Fritz , Daum Gûnther , Zellnig Gûnther
Published: May 2005

Lipid particles of the yeast, Saccharomyces cerevisiae, were isolated to high purity and their components were analysed. The hydrophobic core of this organelle consists of triacylglycerols and steryl esters, which are almost exclusively located to that compartment. Lipid particles are stabilized by a surface membrane consisting of phospholipids and proteins. Electron microscopy confirmed the purity of the preparations and the proposed structure deduced from biochemical experiments. Major proteins of lipid particles have molecular weights of 72, 52, 43 and 34 kDa, respectively. The 43 kDa protein reacts with an antiserum against human apolipoprotein AII. In lipid particles of the yeast mutant strain S. cerevisiae erg6, which is deficient in sterol Δ24‐methyltransferase, this protein is missing thereby identifying the protein and confirming our previous finding (Zinser et al., 1993) that sterol Δ24‐methylation is associated with lipid particles. A possible involvement of surface proteins of lipid particles in the interaction with other organelles is discussed with respect to sterol translocation in yeast.
Isolation and preliminary characterization of Pichia pinus mutants insensitive to glucose repression
Authors: Alamäe Tiina , Simisker Jaan
Published: May 2005

A new method for the isolation of glucose repression‐insensitive mutants in the methylotrophic yeast Pichia pinus was developed. The method is based on screening of small suspension samples derived from 2‐deoxyglucose‐resistant colonies for alcohol oxidase activity. Alcohol oxidase activity was evaluated by determination of formaldehyde excreted by cells. Mutants with glucose non‐repressible alcohol oxidase and catalase synthesis were obtained. All mutants grew poorly on D‐xylose compared to the wild type, whereas growth on L‐arabinose was similar to the wild type. Changes in the glucose transport system were suggested to be responsible for altered growth characteristics and defective glucose repression.
Near‐Stoichiometric interaction between the non‐specific lipid‐transfer protein of the yeast Candida tropicalis and peroxisomal acyl‐coenzyme A oxidase prevents the thermal denaturation of the enzyme in vitro
Authors: Niki Toshiro , Bun‐Ya Masanori , Muro Yoshitaka , Kamiryo Tatsuyuki , Hiraga Yoshikazu
Published: May 2005

A 14‐kDa peroxisomal‐matrix protein, named PXP‐18, of the yeast Candida tropicalis is a structural and functional homologue of the mammalian nonspecific lipid‐transfer protein (identical to sterol carrier protein‐2). PXP‐18 protected acyl‐coenzyme A oxidase (ACO), the rate limiting enzyme of the peroxisomal β‐oxidation of fatty acids, from thermal inactivation at 48°C or 70°C. This effect was dose‐dependent and not replaceable either by chicken egg white lysozyme, which is similar to PXP‐18 (insofar as it is basic, small, and monomeric), or by bovine serum albumin, a carrier of lipids in the blood. ACO was irreversibly denatured by heat treatment at 70°C for 15 min. However, when ACO and PXP‐18 were similarly heat‐treated, they formed a large complex at a molar ratio of PXP‐18 to ACO subunit that was about one, independent of their initial ratio. This near‐stoichiometric complex had ACO activity after a 500‐fold dilution and was accompanied by ACO that was free of PXP‐18 and indistinguishable from native ACO in size and activity. PXP‐18 also protected urate oxidase, another peroxisomal enzyme, from inactivation at 66°C for 15 min and facilitated the renaturation of ACO denatured by 2 M urea. These results indicated that PXP‐18 is active in modulating the structure of peroxisomal enzymes in vitro. It is possible that PXP‐18 functions as a stress protein or as a part of the system that keeps peroxisomal proteins intact.
IX. Yeast sequencing reports. Cloning and sequence of REV7, a gene whose function is required for DNA damage‐induced mutagenesis in Saccharomyces cerevisiae
Authors: Torpey Laurence E. , Gibbs Peter E. M. , Nelson John , Lawrence Christopher W.
Published: May 2005

The function of the REV7 gene is required for DNA damage‐induced mutagenesis in budding yeast, Saccharomyces cerevisiae, and is therefore thought to promote replication past sites of mutagen damage in the DNA template. We have cloned this gene by complementation of the rev7‐2 mutant defect, and determined its sequence. REV7 encodes a predicted protein of Mr 28 759 which is unlike any other protein in the NCBI non‐redundant protein sequence data base, and which is inessential for viability. The sequence of the 3·88 kb yeast genomic fragment containing REV7 has been deposited in Genbank accession number U07228.
II. Yeast sequencing reports. A 12·5 kb fragment of the yeast chromosome II contains two adjacent genes encoding ribosomal proteins and six putative new genes, one of which encodes a putative transcriptional factor
Authors: Démolis Nadine , Mallet Laurent , Jacquet Michel
Published: May 2005

The nucleotide sequence of a 12·5 kb fragment localized to the right arm of chromosome II of Saccharomyces cerevisiae has been determined. The sequence contains eight putative genes. Two of them are contiguous and represent two ribosomal protein genes: SUP46 and URP1. SUP46 is implicated in translation fidelity and encodes the ribosomal protein S13. URP1 is homologous to the rat ribosomal protein gene L21. The open reading frame (ORF) YBR1245 is similar in its N‐terminal part to transcription factors like SRF and MCM1. The ORF YBR1308 shows homology with proteins of the AAA‐family (ATPases Associated with diverse cellular Activities). Two genes are predicted to encode putative membrane proteins. The sequence has been deposited in the EMBL data library under Accession Number U02073.
Consideration of the evolution of the Saccharomyces cerevisiae MEL gene family on the basis of the nucleotide sequences of the genes and their flanking regions
Authors: Turakainen Hilkka , Kristo Paula , Korhola Matti
Published: May 2005

Analysis of the DNA sequences of new members of the Saccharomyces cerevisiae MEL1‐MEL10 gene family showed high homology between the members. The MEL gene family, α‐galactosidase‐coding sequences, have diverged into two groups; one consisting of MEL1 and MEL2 and the other of MEL3‐MEL10. In two S. cerevisiae strains containing five or seven MEL genes each, all the genes are nearly identical, suggesting very rapid distribution of the gene to separate chromosomes. The sequence homology and the abrupt change to sequence heterogeneity at the centromere‐proximal 3′ end of the MEL genes suggest that the distribution of the genes to new chromosomal locations has occurred partly by reciprocal recombination at solo delta sequences.

We identified a new open reading frame sufficient to code for a 554 amino acid long protein of unknown function. The new open reading frame (Accession number Z37509) is located in the 3′ non‐coding region of MEL3‐MEL10 genes in opposite orientation to the MEL genes (Accession numbers Z37508, Z37510, Z37511). Northern analysis of total RNA showed no hybridization to a homologous probe, suggesting that the gene is not expressed efficiently if at all.
The linkage of (1–3)‐β‐glucan to chitin during cell wall assembly in Saccharomyces cerevisiae
Authors: Hartland Robbert P. , Vermeulen Cees A. , Sietsma Johannes H. , Wessels Joseph G. H. , Klis Frans M.
Published: May 2005

Pulse‐chase experiments with [14C]glucose demonstrated that in the cell wall of wild‐type Saccharomyces cerevisiae alkali‐soluble (1–3)‐β‐glucan serves as a precursor for alkali‐insoluble (1–3)‐β‐glucan. The following observations support the notion that the insolubilization of the glucan is caused by linkage to chitin: (i) degradation of chitin by chitinase completely dissolved the glucan, and (ii) disruption of the gene for chitin synthase 3 prevented the formation of alkali‐insoluble glucan. These cells, unable to form a glucan–chitin complex, were highly vulnerable to hypo‐osmotic shock indicating that the linkage of the two polymers significantly contributes to the mechanical strength of the cell wall.

Conversion of alkali‐soluble glucan into alkali‐insoluble glucan occurred both early and late during budding and also in the ts‐mutant cdc24‐1 in the absence of bud formation.
Molecular analysis of the SNF8 gene of Saccharomyces cerevisiae
Authors: Yeghiayan Paula , Tu Jiangling , Vallier Laura G. , Carlson Marian
Published: May 2005

Mutations in the SNF8 gene impair derepresson of the SUC2 gene, encoding invertase, in response to glucose limitation of Saccharomyces cerevisiae. We report here the cloning of the SNF8 gene by complementation. Sequence analysis predicts a 26 936‐dalton product. Disruption of the chromosomal locus caused a five‐fold decrease in invertase derepression, defective growth on raffinose, and a sporulation defect in homozygous diploids. Genetic analysis of the interactions of the snf8 null mutation with spt6/ssn20 and ssn6 suppressors distinguished SNF8 from the groups, SNF1, SNF4 and SNF2, SNF5, SNF6. Notably, the snf8 ssn6 double mutants were extremely sick. Mutations of SNF8 and SNF7 showed similar phenotypes and genetic interactions, and the double mutant combination caused no additional phenotypic impairment. These findings suggest that SNF7 and SNF8 are functionally related. The complete nucleotide sequence of SNF8 has been deposited in GenBank under accession number U10361.
Sequence, mapping and disruption of CCC2, a gene that cross‐complements the Ca2+‐sensitive phenotype of csg1 mutants and encodes a P‐type ATPase belonging to the Cu2+‐ATPase subfamily
Authors: Fu Dadin , Beeler Troy J. , Dunn Teresa M.
Published: May 2005

We have isolated, sequenced, mapped and disrupted a gene, CCC2, from Saccharomyces cerevisiae. This gene displays non‐allelic complementation of the Ca2+‐sensitive phenotype conferred by the csg1 mutation. Analysis of the CCC2p amino acid sequence reveals that it encodes a member of the P‐type ATPase family and is most similar to a subfamily thought to consist of Cu2+ transporters, including the human genes that mutate to cause Wilson disease and Menkes disease. The ability of this gene, in two or more copies, to reverse the csg1 defect suggests that Ca2+‐induced death of csg1 mutant cells is related to Cu2+ metabolism. Cells without CCC2 require increased Cu2+ concentrations for growth. Therefore CCC2p may function to provide Cu2+ to a cellular compartment rather than in removal of excess of Cu2+. The sequence of CCC2 is available through GenBank under accession number L36317.
Transient responses of Candida utilis to oxygen limitation: Regulation of the Kluyver effect for maltose
Authors: Kaliterna Janko , Weusthuis Ruud A. , Castrillo Juan I. , Van Dijken Johannes P. , Pronk Jack T.
Published: May 2005

The facultatively fermentative yeast Candida utilis exhibits the Kluyver effect for maltose: this disaccharide is respired and assimilated but, in contrast to glucose, it cannot be fermented. To study the mechanism of the Kluyver effect, metabolic responses of C. utilis to a transition from aerobic, sugar‐limited growth to oxygen‐limited conditions were studied in chemostat cultures. Unexpectedly, the initial response of maltose‐grown cultures to oxygen limitation was very similar to that of glucose‐grown cultures. In both cases, alcoholic fermentation occurred after a lag phase of 1 h, during which glycerol, pyruvate and D‐lactate were the main fermentation products. After ca. 10 h the behaviour of the maltose‐ and glucose‐grown cultures diverged: ethanol disappeared from the maltose‐grown cultures, whereas fermentation continued in steady‐state, oxygen‐limited cultures grown on glucose. The disappearance of alcoholic fermentation in oxygen‐limited chemostat cultures growing on maltose was not due to a repression of the synthesis of pyruvate decarboxylase and alcohol dehydrogenase. The results demonstrate that the Kluyver effect for maltose in C. utilis does not reflect an intrinsic inability of this yeast to ferment maltose, but is caused by a regulatory phenomenon that affects a key enzyme in maltose metabolism, probably the maltose carrier. The observed kinetics indicate that this regulation occurs at the level of enzyme synthesis rather than via modification of existing enzyme activity.
Nucleotide sequence and characterization of the Saccharomyces cerevisiae RPL19A gene encoding a homolog of the mammalian ribosomal protein l19
Authors: Song Jae Mahn , Cheung Edwin , Rabinowitz Jesse C.
Published: May 2005

A gene designated RPL19A has been identified in the region downstream from the 3′‐end of the Saccharomyces cerevisiae MIS1 gene encoding the mitochondrial C1‐tetrahydrofolate synthase. The gene codes for the yeast ribosomal protein YL19 which exhibits 57·5% identity with the mammalian ribosomal protein L19. RPL19A is one of two functional copies of the YL19 gene located on chromosome II. The disruption of RPL19A has no effect on the growth of the yeast. The RPL19A gene contains an intron located near the 5′‐end. The 5′‐flanking region contains one similar and one complete UASrpg upstream activating sequence. RPL19A was also found to be adjacent to the chromosome II AAC3 gene, encoding the mitochondrial ADP/ATP carrier protein. The nucleotide sequence(s) reported in this paper has been submitted to the GenBanktm/EMBL data bank with the accession number Z36751.
Regulation of carbon metabolism in chemostat cultures of Saccharomyces cerevisiae grown on mixtures of glucose and ethanol
Authors: De Jong‐Gubbels Patricia , Vanrolleghem Peter , Heijnen Sef , Van Dijken Johannes P. , Pronk Jack T.
Published: May 2005

Growth efficiency and regulation of key enzyme activities were studied in carbon‐ and energy‐limited chemostat cultures of Saccharomyces cerevisiae grown on mixtures of glucose and ethanol at a fixed dilution rate. Biomass yields on substrate carbon and oxygen could be adequately described as the net result of growth on the single substrates. Activities of isocitrate lyase and malate synthase were not detected in cell‐free extracts of glucose‐limited cultures. However, both enzymes were present when the ethanol fraction in the reservoir medium exceeded the theoretical minimum above which the glyoxylate cycle is required for anabolic reactions. Fructose‐1,6‐bisphosphatase activity was only detectable at high ethanol fractions in the feed, when activity of this enzyme was required for synthesis of hexose phosphates. Phospho‐enol‐pyruvate‐carboxykinase activity was not detectable in extracts from glucose‐grown cultures and increased with the ethanol fraction in the feed. It is concluded that, during carbon‐limited growth of S. cerevisiae on mixtures of glucose and ethanol, biosynthetic intermediates with three or more carbon atoms are preferentially synthesized from glucose. Synthesis of the key enzymes of gluconeogenesis and the glyoxylate cycle is adapted to the cells′ requirement for these intermediates. The gluconeogenic enzymes and their physiological antagonists (pyruvate kinase, pyruvate carboxylase and phosphofructokinase) were expressed simultaneously at high ethanol fractions in the feed. If futile cycling is prevented under these conditions, this is not primarily achieved by tight control of enzyme synthesis.
Sequence analysis of a 33·1 kb fragment from the left arm of Saccharomyces cerevisiae chromosome X, including putative proteins with leucine zippers, a fungal Zn(II)2‐Cys6 binuclear cluster domain and a putative α2‐SCB‐α2 binding site
Authors: Miosga T. , Schaaff‐Gerstenschläger I. , Chalwatzis N. , Baur A. , Boles E. , Fournier C. , Schmitt S. , Velten C. , Wilhelm N. , Zimmermann F. K.
Published: May 2005

In the framework of the European BIOTECH project for sequencing the Saccharomyces cerevisiae genome, we have determined the nucleotide sequence of the left part of the cosmid clone 232 and the cosmid clone 233 provided by F. Galibert (Rennes Cedex, France). We present here 33,099 base pairs of sequence derived from the left arm of chromosome X of strain S288C. This sequence reveals 17 open reading frames (ORFs) with more than 299 base pairs, including the published sequences for ARG3, LIGTR/LIG1, ORF2, ACT3 and SCP160. Two other ORFs showed similarity with S. cerevisiae genes: one with the CAN1 gene coding for an arginine permease, and one with genes encoding the family of transcriptional activators containing a fungal Zn(II)2‐Cys6 binuclear cluster domain like that found in Ppr1p or Gal4p. Both putative proteins contain a leucine zipper motif, the Can1p homologue has 12 putative membrane‐spanning domains and a putative α2‐SCB‐α2 binding site. In a diploid disruption mutant of ORF J0922 coding for the transcriptional activator homologue, no colonies appeared before 10 days after transformation and then grew slowly. In contrast, haploid disruption mutants showed a growth phenotype like wild‐type cells. One ORF showed weak similarity to the rad4 gene product of Schizosaccharomyces pombe and is essential for yeast growth. Five ORFs showed similarity to putative genes on the right arm of chromosome XI of S. cerevisiae. Two of them have similarity to each other and belong to a family of extracellular proteins that groups mammalian SCP/Tpx‐1, insects Ag3/Ag5, plants PR‐1 and fungi Sc7/Sc14. Three small ORFs are completely contained in the larger ORFs on the corresponding complementary strand and thus probably do not represent real genes. The DNA sequence has been deposited in the EMBL data library under Accession Number X83502.
A regulatedMET3‐GLC7gene fusion provides evidence of a mitotic role forSaccharomyces cerevisiaeprotein phosphatase 1
Authors: Black Sheila , Andrews Paul D. , Sneddon Alan A. , Stark Michael J. R.
Published: May 2005

Saccharomyces cerevisiaepossesses a single essential gene (GLC7) encoding protein phosphatase 1 (PP1). Elevated expression of this gene from theGAL1promoter is highly detrimental to the cell, causing a growth defect and aberrant bud morphology, which leads to cells exhibiting long, extended buds. By comparison, expression ofGLC7from the weakerMET3promoter was without significant effect on either growth or morphology. However, repression ofGLC7expression from theMET3promoter in cells where theMET3‐GLC7fusion was the sole source of PP1 resulted in a mitotic delay. Such cultures showed a massive decrease in the rate of proliferation in conjunction with a significant increase in the proportion of large, budded cells. 4′,6‐diamidino‐2‐phenylindole dihydrochloride (DAPI) staining and anti‐tubulin immunofluorescence analysis of these cells revealed that many were blocked in mitosis, with a short spindle and DAPI‐stained material stretched between the mother and daughter cell within the bud neck. These results support a role for PP1 in the completion of mitosis inS. cerevisiae.
Analysis of a 42·5 kb DNA sequence of chromosome X reveals three tRNA genes and 14 new open reading frames including a gene most probably belonging to the family of ubiquitin–protein ligases
Authors: Huang Meng‐Er , Chuat Jean‐Claude , Galibert Francis
Published: May 2005

We have sequenced a 42,500 bp stretch located on chromosome X of Saccharomyces cerevisiae between the genes MET3 and CDC8. This stretch contains 24 open reading frames (ORFs) of at least 100 amino acids. Ten of these correspond to previously published sequences, whereas of the 14 remaining ORFs, only one, GTD892, has significant similarity to proteins from yeast or other organisms. It may belong to the family of ubiquitin–protein ligases and be involved in the ubiquitin‐dependent proteolytic pathway. In addition, three tRNA genes were recognized, two of which had not been hitherto localized. The sequence has been deposited in the Genome Sequence Data Base under Accession Number L36344.
A short‐chain dehydrogenase gene from Pichia stipitis having D‐arabinitol dehydrogenase activity
Authors: Hallborn Johan , Walfridsson Mats , Penttilä Merja , Keränen Sirkka , Hahn‐hägerdal Bärbel
Published: May 2005

An NAD+‐dependent D‐arabinitol dehydrogenase (polyol dehydrogenase) gene was isolated from Pichia stipitis CBS 6054 and cloned in Saccharomyces cerevisiae. The gene was isolated by screening of a λ‐cDNA library with a zymogram technique. D‐Arabinitol, xylitol, D‐glucitol and galactitol are substrates for the recominant protein. With D‐arabinitol as substrate the reaction product is D‐ribulose. The molecular weight of the native tetramer enzyme is 110 000 Da and the monomer is 30 000 Da. The amino acid sequence is homologous to the short‐chain dehydrogenase family. It is 85·5% identical to a D‐arabinitol dehydrogenase from Candida albicans. The gene in P. stipitis was induced by D‐arabinitol and P. stipitis was able to grow on D‐arabinitol. The physiological role of D‐rabinitol metabolism is discussed.
VII. Yeast sequencing reports. The complete sequence of a 9000 bp fragment of the right arm of Saccharomyces cerevisiae chromosome VII contains four previously unknown open reading frames
Authors: Guerreiro P. , Silva A. Maia E. , Barreiros T. , Rodrigues‐Pousada C. , Arroyo J. , García‐Gonzalez M. , Nombela C. , García‐Saez M. I.
Published: May 2005

We report the sequence of a 9000 bp fragment from the right arm of Saccharomyces cerevisiae chromosome VII. Analysis of the sequence revealed four complete previously unknown open reading frames, which were named G7587, G7589, G7591 and G7594 following standard rules for provisional nomenclature. Outstanding features of some of these proteins were the homology of the putative protein coded by G7589 with proteins involved in transcription regulation and the transmembrane domains predicted in the putative protein coded by G7591. The sequence reported has been deposited in the EMBL data library under Accession Number X82775.
Ribosomal frameshifting in yeast viruses
Authors: Dinman Jonathan D.
Published: May 2005

Proper maintenance of translational reading frame by ribosomes is essential for cell growth and viability. In the last 10 years it has been shown that a number of viruses induce ribosomes to shift reading frame in order to regulate the expression of gene products having enzymatic functions. Studies on ribosomal frameshifting in viruses of yeast have been particularly enlightening. The roles of viral mRNA sequences and secondary structures have been elucidated and a picture of how these interact with host chromosomal gene products is beginning to emerge. The efficiency of ribosomal frameshifting is important for viral particle assembly, and has identified ribosomal frameshifting as a potential target for antiviral agents. The availability of mutants of host chromosomal gene products involved in maintaining the efficiency of ribosomal frameshifting bodes well for the use of yeast in future studies of ribosomal frameshifting.
XIV. Yeast sequencing reports. Sequence analysis of a 30 kb DNA segment from yeast chromosome XIV carrying a ribosomal protein gene cluster, the genes encoding a plasma membrane protein and a subunit of replication factor C, and a novel putative serine/threonine protein kinase gene
Authors: Maurer Kick C. T. , Urbanus Jos H. M. , Planta Rudi J.
Published: May 2005

We have determined the nucleotide sequence of a 30 kb fragment of chromosome XIV of Saccharomyces cerevisiae. The sequence revealed the presence of 19 open reading frames (ORFs) longer than 300 bp. NO422 and NO425 correspond to the split ribosomal protein genes encoding S16A and rp28, respectively, NO450 displays a striking similarity with serine/threonine protein kinase genes, in particular with STE20, and therefore may encode a novel member of this protein family. NO453 is the longest ORF in this DNA segment, having a size of 4908 bp, but its function is not yet known. NO530 encodes the plasma membrane protein Mid1p and NO533 corresponds to the gene coding for a 40 kDa subunit of replication factor C. The remaining ORFs show weak or no homology with proteins in the data bases. The sequence has been submitted to the EMBL data library under Accession Number U23084.
Methylotrophic yeasts as factories for the production of foreign proteins
Authors: Faber Klaas Nico , Harder Wim , Ab Geert , Veenhuis Marten
Published: May 2005

In this contribution we discuss the potential of methylotrophic yeasts as hosts for the high level production of valuable foreign proteins. Recent relevant achievements on the intracellular production or secretion of proteins are summarized. Special attention is paid to a specific advantage of the use of methylotrophic yeasts, namely the possibility of accumulating the foreign gene products inside peroxisomes. This approach may be of major advantage when the protein product is toxic for the host cell and, also, to protect these proteins these proteins from undesired side‐effects such as proteolysis or aggregation.
The low‐affinity component of the glucose transport system in Saccharomyces cerevisiae is not due to passive diffusion
Authors: Gamo Francisco J. , Moreno Eulalia , Lagunas Rosario
Published: May 2005

It has been claimed that the low‐affinity component of glucose transport in Saccharomyces cerevisiae is due to passive diffusion of the sugar across the plasma membrane. We have investigated this possibility. For this purpose we have measured the permeability coefficient of hexoses in this organism. We have found that this coefficient is at least two to three orders of magnitude lower than required to account for the low‐affinity component of glucose transport, and have concluded that this component is not due to passive diffusion.
Yeast sequencing reports. Molecular cloning and sequencing of the hcs gene, which encodes 3‐hydroxy‐3‐methylglutaryl coenzyme a synthase of Schizosaccharomyces pombe
Authors: Katayama Satoshi , Adachi Nami , Takao Kouta , Nakagawa Tsuyoshi , Matsuda Hideyuki , Kawamukai Makoto
Published: May 2005

We have cloned and sequenced the hcs gene, which is thought to encode a 3‐hydroxy‐3‐methylglutaryl coenzyme A (HMG‐CoA) synthase consisting of 447 amino acids, from the fission yeast Schizosaccharomyces pombe. The predicted amino acid sequence of the hcs product of S. pombe has homology with the HMG‐CoA synthase of rat (47·8%), chicken (49·2%), hamster (47·1%) and human cells (46·9%). One of the hcs genes was replaced with a marker gene in the diploid cell. No viable hcs‐disrupted haploid was isolated after tetrad dissection, suggesting that the hcs gene is essential for growth. However the hcs‐defective mutant could be grown on a medium containing 5 mg/ml mevalonate. These results strongly support that the hcs gene encodes HMG‐CoA synthase and S. pombe contains a single copy of the hcs gene. The sequence of the hcs gene has been entered into the public data libraries under Accession Number U32187.
[PSI] and [URE3] as yeast prions
Authors: Wickner Reed B. , Masison Daniel C. , Edskes Herman K.
Published: May 2005

[URE3] is a non‐Mendelian genetic element that mimics recessive mutations in the chromosomal URE2 gene making cells derepressed for nitrogen catabolic enzymes. [PSI] is a non‐Mendelian enhancer of readthrough of translational termination similar in its effects to some mutations in the chromosomal SUP35 gene. Three lines of evidence led to the proposal75 that both [URE3] and [PSI] are prions, infectious proteins analogous to the scrapie agent mediating transmissible spongiform encephalopathies of mammals. (1) Both [PSI] and [URE3] are reversibly curable. (2) [PSI] propagation requires SUP35 and [URE3] propagation requires URE2 with recessive chromosomal mutants having the same phenotypes as the presence of the respective dominant non‐Mendelian element. (3) Overproduction of Sup35p and Ure2p increases the frequency of cells acquiring [PSI] or [URE3], respectively.
Acute heart failure: Clinical presentation, one‐year mortality and prognostic factors
Authors: Rudiger Alain , Harjola Veli‐Pekka , Müller Andreas , Mattila Eero , Säila Petrus , Nieminen Markku , Follath Ferenc
Published: May 2005

Aims

Acute heart failure (HF) is a common but ill‐defined clinical entity. We describe patients hospitalised with acute HF in regard of clinical presentation, mortality, and risk factors for an unfavourable outcome.

Methods and results

We conducted a prospective study including 312 consecutive patients from two European centers hospitalised with acute HF, defined as new onset or worsening of symptoms and signs of HF within 7 days. The mean age was 73 years and 56% were men. Twenty‐eight percent had de‐novo acute HF and 72% a decompensation of chronic HF. Coronary heart disease (CHD) was the most frequent underlying heart disease, elevated blood pressure >150 mmHg and acute ischemia being the most important triggers. Four percent of the patients had cardiogenic shock, 13% presented with pulmonary edema. LV‐EF was <35%, 35−50% and >50% in 35%, 32% and 33% of the patients, respectively. ICU‐treatment was necessary in 39% of the patients. Thirty‐day mortality (11%) was increased in the presence of shock or elevated troponin T levels. Twelve‐month all‐cause mortality (29%) increased in the presence of shock, left ventricular dysfunction, renal insufficiency, CHD, and age.

Conclusions

This prospective study shows that despite modern treatment, morbidity and mortality of patients hospitalised with acute HF remain high.