bluuebunny/crossref_metadata_2025_split huggingface dataset:
RCS1, a gene involved in controlling cell size in Saccharomyces cerevisiae
Authors: Gil Rosario , Zueco Jesús , Sentandreu Rafael , Herrero Enrique 
Published: May 2005

AbstractCloning and sequencing of RCS1, a Saccharomyces cerevisiae gene whose product seems to be involved in timing the budding event of the cell cycle, is described. A haploid strain in which the 3′‐terminal region of the chromosomal copy of the gene has been disrupted produces cells that are, on average, twice the size of cells of the parental strain. The critical size for budding in the mutant is similarly increased, and the disruption mutation is dominant in a diploid heterozygous for the RCS1 gene. Spores from this diploid have a reduced ability to germinate, the effect being more pronounced in the spores carrying the disrupted copy of RCS1. However, disrupted cells recover from α‐factor treatment equally as well as wild‐type cells.
Scarcity of ars sequences isolated in a morphogenesis mutant of the yeast Yarrowia lipolytica
Authors: Fournier Ph. , Guyaneux L. , Chasles M. , Gaillardin C. 
Published: May 2005

AbstractPrevious attempts to isolate autonomously replicating sequences (ars) from the dimorphic yeast Yarrowia lipolytica have been unsuccessful. We isolated a Fil− mutant unable to produce hyphae and growing only in a yeast form to facilitate ars isolation. This mutant was transformed with a Y. lipolytica DNA bank and several unstable clones were obtained. Extrachromosomal plasmids were evidenced in yeast recovered in Escherichia coli and characterized by restriction mapping. They were able to retransform Fil− and Fil+ yeast strains at high frequency and transformants displayed a slightly unstable phenotype. The detailed analysis of the plasmids showed that only two different ars sequences had been isolated, each of them corresponding to a unique sequence in the Y. lipolytica genome. We concluded that functional ars sequences that can be cloned on plasmids are rare in this yeast.
On the dependence of spontaneous mutation rates on the functional state of genes
Authors: Korogodin V. I. , Korogodina V. L. , Fajszi Cs. , Chepurnoy A. I. , Mikhova‐Tsenova N. , Simonyan N. V. 
Published: May 2005

AbstractSpontaneous mutation of some genes was studied in haploid adenine and leucine auxotrophic yeast Saccharomyces.It was shown that a decrease in the amount of adenine (from 500 to 0 mgl−1) or leucine (from 300 to 0·3 mgl−1) in the medium, simultaneously with the transition from repression to derepression of the biosynthesis of these metabolites, resulted in a 15‐ to 150‐fold increase in the reversion rate of genes ade2 and leu2, respectively, for different strains. At the same time the mutation rate of suppressor genes varied relatively little (up to five‐fold), and that of gene lys1 did not change at all. It was also demonstrated (on gene leu2) that the mutation rate is determined by the composition of the nutrient medium at the time of the S‐phase of the cell cycle and it does not depend on the cultivation conditions during the presynthetic period.We discuss the hypothesis that derepressed genes mutate with a significantly higher rate than genes in the repressed state.
Quantitation of readthrough of termination codons in yeast using a novel gene fusion assay
Authors: Firoozan Mandy , Grant Christopher M. , Duarte Julio A. B. , Tuite Mick F. 
Published: May 2005

AbstractA simple quantitative in vivo assay has been developed for measuring the efficiency of translation of one or other of the three termination codons, UAA, UAG and UGA in Saccharomyces cerevisiae. The assay employs a 3‐phosphoglycerate kinase‐β‐galactosidase gene fusion, carried on a multicopy plasmid, in which the otherwise retained reading frame is distupted by one or other of the three termination codons. Termination readthrough is thus quantitated by measuring β‐galactosidase in transformed strains. Using these plasmids to quantitate the endogenous levels of termination readthrough we show that readthrough of all three codons can be detected in a non‐suppressor (sup+) strain of S. cerevisiae. The efficiency of this endogenous readthrough is much higher in a [psi+] strain than in a [psi−] strain with the UGA codon being the leakiest in the nucleotide context used. The utility of the assay plasmids for studying genetic modifiers of nonsense suppressors is also shown by their use to demonstrate that the cytoplasmic genetic determinant [pse+] broadens the decoding properties of a serine‐inserting UAA suppressor tRNA (SUQ5) to allow it to translate the other two termination codons in the order of efficiency UAA > UAG > UGA.
Calcium‐dependent secretory vesicle‐binding and lipid‐binding proteins of Saccharomyces cerevisiase
Authors: Creutz Carl E. , Snyder Sandara L. , Kambouris Nicholas G. 
Published: May 2005

AbstractYeast (Saccharomyces cerevisiae) cytosol was examined for the presence of calcium‐dependent membrane‐ or lipidbinding proteins that might paly fundamental roles in membrane‐associated phenomena in stimulated cells. A complex group of proteins was isolated from late log phase cultures of yeast strain YP3 on the basis of calcium‐dependent association with yeast secretory vesicles isolated from the temperature‐sensitive sec6‐4 secretory mutant. The masses of the major proteins in this group were 32, 35, 47, 51, 55, 60, and 120 kDa. A similar group of proteins was isolated by calcium‐dependent association with bovine brain lipids enriched in the predominant acidic phospholipids of the yeast secretory vesicles. The 47 kDa protein was highly purified when commerical yeast cake was used as the source of yeast cytosol. The 32 kDa and 60 kDa proteins were demonstrated to reassociate with lipids at calcium concentrations of 100 μM or higher, while no association was promoted by 2 mM‐magnesium. The 47 kDa protein could be removed from lipids by reducing the calcium concentration to between 1 and 32 μM. The sequences of peptides isolated from digests of several of these proteins indicate that they are novel proteins but are insufficient to judge the possible homology of these proteins with mammalian membrane‐binding proteins. The sequence data may be adequeate to permit isolation and modification of the corresponding genes in order to assess the possible funtion of this class of proteins in stimulated cells.
Simultaneous expression of the S and L surface antigens of hepatitis B, and formation of mixed particles in the methylotrophic yeast, Hansenula polymorpha
Authors: Janowicz Zbigniew A. , Melber Karl , Merckelbach Armin , Jacobs Eric , Harford Nigel , Comberbach Martin , Hollenberg Cornelis P. 
Published: May 2005

AbstractAn expression system has been developed for the methylotrophic yeast Hansenula polymorpha and used to co‐express both the L (preS1‐S2‐S) and S hepatitis B surface antigens (HBsAg) under the control of strong methanol‐inducible promoters derived from the methanol oxidase and from the formate dehydrogenase genes. A unique feature of this H. polymorpha expression system is the possibility of integrating up to 100 copies of an expression cassette via a multimeric integration mechanism. Several multimeric integrants containing various numbers of L and S expression cassettes were constructed to give a spectrum of strains characterized by different L to S ratios. The expression level of S antigen was 5–8% of the total soluble cell protein. Analysis by sucrose and CsCl density gradient centrifugation and by particle‐specific immunoassays demonstrated that the synthesized HBsAg spontaneously assembled into composite subviral particles containing both S and L proteins. Only a minor portion of the L protein was found to be glycosylated. These H. polymorpha‐derived composite particles can be used for the production of a hepatitis B virus vaccine with the potential for improved immunogenicity due to the presence of a wider spectrum of epitopes and negligible glycosylation.
Expression of the α‐galactosidase from Cyamopsis tetragonoloba (guar) by Hansenula polymorpha
Authors: Fellinger Arthur J. , Verbakel John M. A. , Veale Rosemary A. , Sudbery Peter E. , Bom Isaac J. , Overbeeke Nico , Verrips C. Theo 
Published: May 2005

AbstractThe methylotrophic yeast Hansenula polymorpha, a host organism for the production of heterologous proteins, has been applied to produce the α‐galactosidase from the plant Cyamopsis tetragonoloba (guar). The yeast/Escherichia coli shuttle expression vector used is based on the origin of replication of the endogenous 2 μm plasmid of Saccharomyces cerevisiae and the LEU2 gene of S. cerevisiae for selection in H. polymorpha. In the expression vector, the α‐galactosidase is controlled by the methanol‐regulated promoter from the methanol oxidase gene, MOX, of H. polymorpha. The signal sequence of SUC2 (invertase) from the yeast S. cerevisiae, was used to ensure secretion of the α‐galactosidase enzyme. After transformation and stabilization, the expression vector was stably integrated in the genome. The active α‐galactosidase enzyme was efficiently secreted (>85%) and after methanol induction, the expression level was 42 mg/l. Amino‐terminal sequencing of the purified α‐galactosidase enzyme synthesized by H. polymorpha showed that the S. cerevisiae invertase signal sequence was correctly processed by H. polymorpha. The secreted α‐galactosidase was glycosylated and had a sugar content of 9·5%. The specific activity of the α‐galactosidase produced by H. polymorpha was 38 U mg−1 compared to 100 U mg−1 for guar α‐galactosidase. Deglycosylation of the H. polymorpha α‐galactosidase restored the specific activity completely.
The role of pyruvate decarboxylase in the Kluyver effect in the food yeast, Candida utilis
Authors: Sims A. P. , Stålbrand H. , Barnett J. A. 
Published: May 2005

AbstractThe glucose‐fermenting yeast, Candida utilis cannot use the β‐D‐glucoside, cellobiose, anaerobically, although it is able to do so aerobically. β‐Glucoside transport and hydrolysis and pyruvate decarboxylase activities of this yeast were measured aerobically and anaerobically. β‐Glucoside transport was five‐fold faster aerobically than anaerobically, but there was no corresponding difference in β‐glucosidase activity. Pyruvate decarboxylase activity varied greatly, being synthesized de novo in response to the presence of D‐glucose and anaerobic conditions and about 50% deactivated on the removal of D‐glucose or the addition of air. Activation and deactivation were rapidly reversible. Failure to utilize cellobiose anaerobically, in particular, and the Kluyver effect, in general, probably depends on much reduced glycolytic flux, associated under anaerobic conditions, with (i) lower transport rate, (ii) low substrate affinity of the relevant glycosidase and (iii) deactivation of pyruvate decarboxylase. So, in addition to the complex effects of oxygen, anaerobiosis and specific sugars on induction, repression and derepression, there are fine controls on pyruvate decarboxylase activity, leading to fast activation or deactivation of the enzyme.
TDH2 is linked to MET3 on chromosome X of Saccharomyces cerevisiae
Authors: Mountain Harry A. , Korch Christopher 
Published: May 2005

AbstractThe MET3 gene of Saccharomyces cerevisiae was cloned and its restriction map was found to differ in the upstream region from an earlier published map (Cherest et al. Gene 34, 269–281, 1985) and nucleotide sequence (Cherest et al. Mol. Gen. Genet. 210, 307–313, 1987). Southern blot analysis of genomic DNA from strains S288C and FL100 (the genetic backgrounds from which these different copies of the gene had been cloned) showed that our clone from a S288C‐based library had the same restriction map as the chromosomal DNA from both of the strains. Comarison of the nucleotide sequence of the two clones indicated that the earlier published clone probably represented a cloning artifact. In our clone, we found upstream of MET3, the nucleotide sequence of the TDH2 gene (Holland and Holland, J. Biol. Chem. 255, 2596–2605, 1980). The chromosomal orientation of the two genes was determined to be MET3–TDH2–CEN10.
The sequence of a 6·3 kb segment of yeast chromosome III reveals an open reading frame coding for a putative mismatch binding protein
Authors: Valle Giorgio , Bergantino Elisabetta , Lanfranchi Gerolamo , Carignani Giovanna 
Published: May 2005

AbstractWe report the sequence of a 6·3 kb segment of DNA mapping near the end of the right arm of chromosome III of Saccharomyces cerevisiae. The sequence reveals a major open reading frame coding for a putative protein of 1047 amino acids with a striking similarity to the bacterial proteins involved in recognition of mismatched DNA base pairs. This is particularly interesting as the existence of a yeast mismatch repair system similar to that of bacteria has been postulated for some years, but a yeast protein homologous to the bacterial mismatch binding protein had not been identified.The results of a comparison of the putative yeast mismatch binding protein with the bacterial mismatch binding proteins and with two cognate mammalian sequences, support the idea that a similar mismatch repair system may be present also in mammalian cells. The possibility that all of these proteins may have evolved from a common ancestral gene is also discussed.
Yeast flocculation: Reconciliation of physiological and genetic viewpoints
Authors: Stratford Malcolm 
Published: May 2005

AbstractYeast floccultion results from surface expression of specific proteins (lectins). Two flocculation phenotypes were suggested by physiological and biochemical tests, whereas genetic data suggested a larger number of mechanisms of flocculation. After reviewing the biochemistry, physiology and genetics of flocculation, a new hypothesis combining the data available from these different sources, is proposed.Flocculation results when lectins present on flocculent cell bind sugar residues of neighbouring cell walls. These sugar receptors are intrinsic to the mannan comprizing cell walls of Saccharomyces cerevisiae. Two lectin phenotypes were revealed by sugar inhibition studies. The gluco‐ and mannospecific NewFlo phenotype is not, as yet, found in genetically defined strains. Mannospecific flocculation (Flo 1 phenotype) is found in strains containing the genes FLO1, FLO5 and FLO8. This phenotype is also found following mutation of the TUP1 or CYC8 loci, in previously non‐flocculent strains. It is therefore proposed that the structure gene for mannospecific flocculation is common or possibly unbiquitous in non‐flocculent strains and in consequence, FLO1, FLO5 and FLO8 are probably regulatory genes, exerting positive control over the structure gene.Flocculation expression requires lectin secretion to the cell surface. Many of the observed ‘suppressions’ of flocculation may be due to mutations of the secretory process, involved in transporting structural proteins to the cell wall.The possible involvement of killer L double‐stranded RNA with flocculation is suggested, given the lectin properties of viral coat proteins nad an association between L double‐stranded RNA and the Flo 1 phenotype.
The small heat‐shock protein Hsp26 of Saccharomyces cerevisiae assembles into a high molecular weight aggregate
Authors: Bentley Nicola J. , Fitch Ian T. , Tuite Mick F. 
Published: May 2005

AbstractHsp26 is one the major small heat‐shock proteins (Hsp) of the Saccharomyces cerevisiae. yet its cellular role remains to be discovered. To examine the cellular consequences of overexpression of Hsp26, the gene encoding this protein (HSP26) was overexpressed from a multicopy plasmid using either its own promoter or by coupling it to the effecient constitutive PGK promoter. The PGK promoter provided the opportunity to overexpress Hsp26 under nonstress conditions and such high level synthesis, prior to a lethal heat shock (50°C), gave a small but reproducible elevation in thermotolerance. In transformed strains overexpressing Hsp26 under either stressed or non‐stress conditions, the Hsp26 polypeptide was recovered almost exclusively as a high molecular weight aggregate. This high molecular weight aggregate (or heat‐shock granule, HSG) was purified by differential centrifugation and sucrose gradient density centrifugation and shown, by electron microscopic analysis, to be of a uniform size (15–25 nm diameter). Analysis of the purified HSG demonstrated that it had a molecular weight of 550 kDa, yet contained no other integral polypeptides or other macromolecules.
Sequence of a 10·7 kb segment of yeast chromosome XI identifies the APN1 and the BAF1 loci and reveals one tRNA gene and several new open reading frames including homologs to RAD2 and kinases
Authors: Jacquier Alain , Legrain Pierre , Dujon Bernard 
Published: May 2005

AbstractWe report here the DNA sequence of a segment of chromosome XI of Saccharomyces cerevisiae extending over 10·7 kb. The sequence was determined using a double‐strand sequencing strategy adapted from the random‐clone strategy. The segment contaiins seven non‐overlapping long open reading frames, YKL500, 505, 510, 513, 516, 518 and 520 and one tRNA gene. YKL505 and YKL513 are two already sequenced genes, the BAF1/ABF1 and the APN1 genes, respectively. YKL510 exhibits a strong homology to the RAD2 protein and YKL516 is presumabloy a protein kinase.
Identification of a novel secreted glycoprotein of the yeast Saccharomyces cerevisiae stimulated by heat shock
Authors: Lupashin V. V. , Kononova S. V. , Ratner Ye. N. , Tsiomenko A. B. , Kulaev I. S. 
Published: May 2005

AbstractA new secreted yeast glycoprotein with an Mr of about 400 kDa (gp400) has been found. The glycoprotein is an O‐mannosylated oligomer, whose synthesis and export into culture medium are stimulated by heat shock. Intracellular transport of gp400 is carried out by membrane vesicles distinct form the known constitutive scretory vesicles. Immunological analysis revealed gp400 only in Saccharomyces species.
Molecular cloning and physical analysis of an 8·2 kb segment of chromosome XI of Saccharomyces cerevisiae reveals five tightly linked genes
Authors: Abraham Philip R. , Mulder Anita , Riet Jan Van'T , Planta Rudi J. , Raué Hendrik A. 
Published: May 2005

AbstractThe nucleotide sequence of 6472 base pairs of an 8·2 kb segment of Saccharomyces cerevisiae chromosome XI has been determined. The sequence contains a cluster of four long open reading frame (ORF) designated YKL2, YKL3, YKL4 and TGL1 in the same orientation, flanked at the 5′‐end by a divergent incomplete ORF (YKL1). Transcription and Southern analyssis of the four complete ORFs showed that all are expressed and are present in single copy on the haploid genome. The average codon adaption index of the coding regions is approximately 0·2, suggesting that these genes are lowly expressed. The upstream regions of all four genes as well as the YKL1 ORF contain putative promoter elements previously found to be characteristic of nuclear genes encoding mitochondrial proteins. Significant sequence similarities were found between the YKL3 protein and Escherichia coli ribosomal protein S2 as well as between the TGL1 protein and triglyceride lipases from rat salivary gland and human gastric tissue. The 3′‐end of the 6472 bp nucleotide sequence overlaps with the upstream region of the previously identified CTK1 gene, encoding the largest subunit of CTD kinase (Lee, J. M. and Greenleaf, A. L., 1991, Gene Expression 2, 149–167), thereby increasing the number of genes on the 8·2 kb fragment to at least five. The transcripts of these genes represent approximately 83% of the DNA fragment, making it one of the most highly transcribed regions of the yeast chromosome analysed to date.
Molecular cloning and analysis of autonomous replicating sequence of Candida maltosa
Authors: Sasnauskas Kestutis , Jomantienè Rasa , Lebedienè Edita , Lebedys Jurgis , Januška Albinas , Janulaitis Arvydas 
Published: May 2005

AbstractA Candida maltosa chromosomal DNA fragment which confers high frequency transformation of C. maltosa and autonomous replication of recombinant plasmids was cloned and sequenced. Analysis of the nucleotide sequence of the cloned DNA revealed a sequence homologous for C. maltosa autonomously replicating sequence (ARS) elements. Vector pRJ1 for C. maltosa was constructed, which contained a 1.3 kb ARS sequence, pICEM‐19H and the ADE1 gene of C. maltosa. Southern blot analysis suggested that the copy number of pRJ1 in C. maltosa was approximately 20 per genome. The sequence analysis also revealed an open reading frame, encoding a polypeptide with high homology (70%) to the RS15 protein of Brugia pagangi. This open reading frame has an intron with canonical sites for correct splicing in Saccharomyces cerevisiae.
N‐linked glycosylation of proteinase B precursors of the yeast Saccharomyces cerevisiae is not require for proper targeting or processing of the enzyme
Authors: Nebes Vicki L. , Jones Elizabeth W. 
Published: May 2005

AbstractProteinase B precursors are modified by an N‐linked carbohydrate side chain at Asn 314. Glycosylation at this position is not required for proper localization, processing, or activation of the enzyme.
SEC6 encodes an 85 kDa soluble protein required for exocytosis in yeast
Authors: Potenza Marc , Bowser Robert , Müller Heike , Novick Peter 
Published: May 2005

AbstractThe SEC6 gene encodes a protein required for an event leading to fusion of post‐Golgi vesicles with the plasma membrane in Saccharomyces cerevisiae cells. The gene was cloned by complementation of the temperature‐sensitive growth defect of a sec6‐4 strain. The nucleotide sequence was determined and the longest open reading frame was found to encode an 85 kDa protein of 733 amino acids. The Sec6 protein is predicted to be hydrophilic and is found predominantly in the soluble fraction of a yeast lysate, in a species that sediments with a coefficient of 14S. No extnsive homology was found with known proteins of the database. Gene disruption and marker rescue experiments indicate that SEC6 is a single copy gene essential for growth. Overproduction of Sec6p does not suppress any of the other lateactind sec mutants, yet sec6‐4 does display synthetic lethality with sec8‐9, suggesting that the two products may fulfill inter‐related functions.
Development of the yeast Pichia pastoris as a model organism for a genetic and molecular analysis of peroxisome assembly
Authors: Gould Stephen J. , McCollum Dannel , Spong Allan P. , Heyman John A. , Subramani Suresh 
Published: May 2005

AbstractWe describe the isolation of mutants of the yeast Pichia pastoris that are deficient in peroxisome assembly (pas). These mutants of P. pastoris can be identified solely by their inability to grow on methanol and oleic acid, the utilization of which requires peroxisomal enzymes, and are defined by the absence of normal peroxisomes as judged by electron microscopy and biochemical fractionation experiments. These mutants are the result of genetic defects at single loci and represent at least eight different complementation groups. The isolation of pas mutants of P. pastoris by a simple screen for mutants unable to use methanol and oleic acid represents a significantly more efficient method for identification of pas mutants than is possible in other organisms. To exploit this advantage fully we also developed new reagents for the genetic and molecular manipulation of P. pastoris. These include a set of auxotropic strains with an essentialiy wild type genetic background, plasmids that act as Escherichia coli–P. pastoris shuttle vectors, and genomic DNA libraries for isolation of P. pastoris genes by functional complementation of mutants or by nucleic acid hybridization. The availability of numerous pas mutants and the reagents necessary for their molecular analysis should lead to the isolation and characterization of genes involved in peroxisome assembly.