bluuebunny/crossref_metadata_2025_split huggingface dataset:
Known heat‐shock proteins are not responsible for stress‐induced rapid degradation of ribosomal protein mRNAs in yeast
Authors: Galego Lisete , Barahona Isabel , Alves Ana‐Paula , Rodrigues‐Pousada Claudina , Vreken Peter , Raué Hendrik A. , Planta Rudi J. 
Published: May 2005

AbstractWe have previously shown that the heat‐induced enhanced decay of yeast mRNAs encoding ribosomal proteins (rp‐mRNAs) requires ongoing transcription during the heat treatment [Herruer et al. (1988) Nucl. Acids Res. 16, 7917]. In order to determine whether this requirement reflects the need for heat‐shock protein (hsp), we analysed the effect of heat shock on rp‐mRNA levels in several yeast strains in which each of the heat‐shock genes encoding hsp26, hsp35 or hsp83 had been individually disrupted. In all three strains we still observed increased degradation of rp‐mRNAs immediately after the temperature shift, demonstrating that hsp26, hsp35 and hsp83 are not required for this effect. Accelerated turnover of rp‐mRNA was also found to occur upon raising the growth temperature of a mutant strain that contains a disruption of the gene specifying the heat‐shock transcription factor and in wild‐type yeast cells treated with canavanine, an arginine analogue that will be incorporated into all known hsps and that is known to cause misfolding of the polypeptide chain. Latter observation suggests that enhanced rp‐mRNA decay is a more general stress‐related phenomenon. Taken together, these data strongly indicate that the trans‐acting factor required for the increase in the rate of degradation of rp‐mRNAs upon stress is not one of the known yeast hsps.
Molecular analysis of HEM6 (HEM12) in Saccharomyces cerevisiae, the gene for uroporphyrinogen decarboxylase
Authors: Diflumeri Celestino , Larocque Robert , Keng Teresa 
Published: May 2005

AbstractHEM6 (HEM12) in Saccharomyces cerevisiae encodes uroporphyrinogen decarboxylase, the fifth enzyme in the heme biosynthetic pathway. The HEM6 (HEM12) gene was cloned by complementation of heme auxotrophy of a hem6 mutant. Sequence analysis revealed an open reading frame of 1086 nucleotides. The predicted amino acid sequence of HEM6 (HEM12) shows extensive homology to those reported for uroporphyrinogen decarboxylase from mammalian sources. Expression of HEM6 (HEM12) was investigated and was found to increase two‐fold in a non‐fermentable carbon source. However, HEM6 (HEM12) transcription was unaffected by heme or by intermediates in the heme biosynthetic pathway. In addition, HEM6 (HEM12) expression is not regulated by the transcriptional activator complex HAP2–3–4, as has been shown for some genes encoding heme biosynthetic enzymes.
Cloning and sequencing of the URA3 gene of Kluyveromyces marxianus CBS 6556
Authors: Bergkamp Ronald J. M. , Geerse Ruud H. , Planta Rudi J. , Verbakel John M. A. 
Published: May 2005

AbstractThe URA3 gene, coding for orotidine‐5′‐phosphate decarboxylase, from Kluyveromyces marxianus CBS 6556, was isolated from a genomic DNA library. The K. marxianus URA3 gene encodes a protein of 267 amino acids with a calculated molecular weight of 29·3 kDa. Comparison of the K. marxianus protein with the corresponding enzymes of Saccharomyces cerevisiae and Kluyveromyces lactis showed amino acid sequence identifies of 81% and 88%, respectively. Using contour‐clamped homogeneous electric field gel electrophoresis, the genomic copy was found to be located on chromosome VI. We have used the cloned gene for the construction of a K. marxianus leu2 mutant. This mutant contains no heterologous sequences, which is essential to make it acceptable for application in the food industry.
Isolation of single yeast cells by optical trapping
Authors: Grimbergen Jos A. , Visscher Koen , De Mesquita Daniel S. Gomes , Brakenhoff G. J. 
Published: May 2005

AbstractIndividual yeast cells can be successfully isolated and recultured on plates with a new isolation method making use of optical trapping with infrared laser light. The cells can be selected on morphological criteria by high resolution microscopy. The isolation device is constructed from two coverslips separated by spacers, in which selected cells are transferred to a plastic capillary, using the optical trap. To test the procedure, selection experiments were done with a mixture of two Saccharomyces cerevisiae strains, distinguishable both in fluorescence microscopy and on agar plates. These experiments showed that only selected cells were isolated, and close to 100% of the isolated stationary‐phase cells formed colonies on agar plates, indicating a high recovery. A lower recovery was obtained with exponential‐phase cells, possibly because of a higher sensitivity to laser irradiation. Applications for this method may include the isolation of mutants with altered morphology and the isolation of subpopulations of yeast cultures, for their separate investigation or for the initiation of pure cultures.
Different signals control the activation of glycolysis in the yeast Saccharomyces cerevisiae
Authors: Boles Eckhard , Zimmermann Friedrich K. , Heinisch Jürgen 
Published: May 2005

AbstractThe glycolytic pathway in Saccharomyces cerevisiae is activated by fermentable sugars at several steps. Mutants with deletions of genes coding for enzymes of the upper part of glycolysis were used to characterize the triggering mechanisms. Synthesis of fructose‐2,6‐bisphophate is catalysed by two 6‐phosphofructo‐2‐kinase isoenzymes, one of which is activated by fermentable sugars while synthesis of the second enzyme is induced (Kretschmer and Fraenkel, 1991). Increase in the level of fructose‐2, 6‐bisphosphate is demonstrated to depend on an internal metabolite upstream of the phosphoglucose isomerase reaction. The signalling process correlates with distinct temporal changes in the concentration of glucose‐6‐phosphate but not with its absolute level, indicating an adaptational mechanism. It is independent of the uptake and phosphorylation systems used by different sugars. Interestingly, this increase, although delayed, could also be observed in strains lacking the rapid cAMP increase after sugar addition which is thought to be responsible for the activating process. Synthesis of glucose‐6‐P and fructose‐6‐P is needed for the complete induction of pyruvate kinase and inactivation of fructose‐1,6‐bisphosphatase. On the other hand, induction of pyruvate decarboxylase depends mainly on a signal in the lower part of glycolysis.
The multifunctional regulatory proteins ABF1 and CPF1 are involved in the formation of a nuclease‐hypersensitive region in the promoter of the QCR8 gene
Authors: De Winde Johannes H. , van Leeuwen Hans C. , Grivell Leslie A. 
Published: May 2005

AbstractThe abundant DNA‐binding proteins ABF1 and CPF1 are members of a family of global regulators with diverse chromosomal functions in the yeast Saccharomyces cerevisiae. Recent evidence suggests that these protein factors may be involved in establishing and maintaining well‐defined chromatin structures in promoter regions and other genetic elements. We have investigated the involvement of ABF1 and CPF1 in chromatin organization at the QCR8 gene, encoding subunit VIII of the mitochondrial ubiquinol‐cytochrome c oxidoreductase. The promoter region of the QCR8 gene contains overlapping binding sites for ABF1 and CPF1. Nucleosome positioning studies indicate that the QCR8 gene is associated with a phased array of nucleosomes under both catabolite‐repressed and derepressed growth conditions. Analysis of binding site mutants reveals that both ABF1 and CPF1 are involved in maintaining a nuclease‐hypersensitive region in the QCR8 promoter. The chromatin structure at QCR8 during steady‐state growth is, however, mainly dependent on binding of ABF1 to the promoter region. Implications of these findings for the role played by ABF1 and CPF1 in the regulation of mitochondrial biogenesis and other processes important for cell growth and division will be discussed.
Cloning and characterization of the SEC18 gene from Candida albicans
Authors: Nieto Almudena , Sentandreu Rafael , Agudo Lucas Del Castillo , Sanz Pascual 
Published: May 2005

AbstractThe SEC18 gene product is required for protein transport at different stages in the Saccharomyces cerevisiae secretory pathway. The homologous SEC18 gene from Candida albicans has been cloned by complementation of a sec18‐1 S. cerevisiae thermosensitive mutant using a C. albicans genomic library in YRp7. Sequence analysis of the gene revealed a 2382‐bp open reading frame which coded for a protein of 88 926 kDa. By an in vitro transcription–translation coupled reaction of the C. albicans SEC18 gene, a protein of approximately 85 kDa was obtained. Hydrophobicity analysis of the protein did not show any predicted signal sequence nor transmembrane anchor domain. These results and the fact that glycosylation was absent in the protein indicated that C. albicans Sec18p did not enter in the secretory pathway. The alignment of the amino acid sequence revealed that the SEC18 gene from C. albicans was homologous to the SEC18 from S. cerevisiae (50% amino acid identity) and to the gene that coded the N‐ethylmaleimide‐sensitive factor (NSF) protein (43% amino acid identity). Moreover, the C. albicans Sec18p also showed the putative ATP binding site present in S. cerevisiae Sec18p and in NSF.
Isolation and DNA sequence of the STE14 gene encoding farnesyl cysteine: Carboxyl methyltransferase
Authors: Ashby Matthew N. , Errada Patrick R. , Boyartchuk Victor L. , Rine Jasper 
Published: May 2005

AbstractWe isolated a mutant defective in C‐terminal farnesyl cysteine:carboxyl methyltransferase activity from a screen for mutations causing a‐specific sterility. A genomic fragment was cloned from a yeast multi‐copy library that restored mating. Both the cloned gene and the sterile mutation were allelic to the STE14 gene. A ste14‐complementing 2·17 kb BamHI fragment subclone was sequenced and found to encode a 239 amino acid protein with a molecular weight of 27,887 Daltons. The hydrophobicity profile of the methyltransferase reveals the presence of at least five potential transmembrane domains. In comparisons of the C‐terminal methyltransferase amino acid sequence with those in the PIR and Swiss protein databases, no significantly similar sequences were found nor were conserved regions from other methyltransferases present.
KTR2: A new member of the KRE2 mannosyltransferase gene family
Authors: Lussier Marc , Camirand Anne , Sdicu Anne‐Marie , Bussey Howard 
Published: May 2005

AbstractThe KTR2 gene from Saccharomyces cerevisiae was identified by polymerase chain reaction amplification of genomic DNA using primers derived from regions of high homology between the products of three yeast genes, KRE2, YUR1 and KTR1. The product encoded by the KTR2 gene is a predicted type II membrane protein of 425 amino acid residues with a short cytoplasmic N‐terminus, a membrane‐spanning region and a large lumenal domain containing four potential N‐glycosylation sites. Ktr2p has 58% identity with Yur1p, 39% with Ktr1p and 34% with Kre2p. One member of this gene family, KRE2 (also known as MNT1; Häusler and Robbins, 1992), encodes an α‐1,2 mannosyltransferase which adds the third mannose onto O‐linked glycoprotein side‐chains (Häusler et al., 1992). In contrast to KRE2 null mutants, which produce shortened (two‐mannose) chains, mutants harboring a KTR2 gene disruption synthesize O‐linked chains with the wild‐type patterns of five mannose residues. A null mutation in KTR2 leads to partial resistance to killer toxin and hints that KTR2, which encodes a putative mannosyltransferase, is involved in extracellular matrix assembly.
Regulation of the amino acid permeases in nitrogen‐limited continuous cultures of the yeast Saccharomyces cerevisiae
Authors: Olivera Hiram , González Alicia , Peña Antonio 
Published: May 2005

AbstractIn the yeast Saccharomyces cerevisiae, there is a general amino acid permease, regulated by nitrogen catabolite repression, and several specific permeases whose nitrogen regulation is not well understood. In this study, we used continuous cultures to analyse the effect of nitrogen limitation and pH on the activity of general and several specific amino acid permeases. General permease activity was maximal in severe nitrogen limitation and diminished 400‐fold in cells grown under nitrogen excess. For the specific permeases, the maximal uptake activity was found between mild limitation and nitrogen excess, while very small activity was detected under strict limitation. These results indicate that the nitrogen regulation of the general and the specific amino acid carriers is coordinated in such a way that no redundancy exists in amino acid transport. The regulation of the specific permeases was similar to that found for a system with anabolic function in nitrogen metabolism.All of these permeases are supposed to work through a proton symport mechanism, and thus rely on pH gradients to carry out their function. We studied the effect of pH on the kinetic constants of the general permease. Our results show that the effect of pH on the Km was different for acidic, neutral and basic amino acids, while the effect on Vmax was independent of the electrical charge of the amino acids.
PFK2, ISP42, ERG2 and RAD14 are located on the right arm of chromosome XIII
Authors: Heinisch Jûrgen J. 
Published: May 2005

AbstractAn 11 kb yeast DNA insertion isolated from a genomic library by complementation of a phosphofructokinase‐deficient strain was used as a source for a partial sequence analysis. Four genes were shown to reside on this fragment, namely PFK2, ISP42, ERG2 and RAD14. PFK2 was mapped previously to the right arm of chromosome XIII, locating the latter three genes to the same chromosome.
The RHO4a and NUD1 genes on Saccharomyces cerevisiae chromosome XI
Authors: Van Vliet‐Reedijk Joke C. , Planta Rudi J. 
Published: May 2005

AbstractSequence analysis of a 4 kb fragment from the right arm of Saccharomyces cerevisiae chromosome XI, in combination with Northern hybridization experiments revealed the presence of two genes, designated RHO4a and NUD1. The first gene encodes a 32 kDa protein showing significant sequence similarity with members of the ras family. Its 3′‐terminal sequence is virtually identical to a sequence published previously as the RHO4 gene [Matsui and Toh‐e, Gene 114 (1992), 43–49], which, however, appears to start at an internal ATG codon. The RHO4a sequence also overlaps the 5′‐terminal sequence of the RNC1 gene [Chow et al., Nucl. Acids Res. 20 (1992), 5215–5221] proposed to encode the yeast yNucR endo/exonuclease. The remainder of this RNC1 gene overlaps with the 5′‐end of the NUD1 gene. However, the RNC1 sequence lacks a portion of 276 bp that in our fragment is part of the intergenic region separating RHO4a and NUD1. From these results we conclude that the proposed RNC1 gene is the result of a cloning artefact and that the yNucR protein is instead encoded by the NUD1 gene.
Nucleotide sequence analysis of the 5·8S rDNA and adjacent ITS2 region of Candida albicans and related species
Authors: Lott Timothy J. , Kuykendall Randall J. , Reiss Errol 
Published: May 2005

AbstractWe have determined the nucleotide sequence for the DNA encoding the 5·8S RNAs and downstream internal transcribed spacer (ITS2) regions for Candida albicans and the taxonomically related species C. parapsilosis, C. tropicalis, C. glabrata and C. krusei. Phylogenetic analysis of all known fungal 5·8S RNA sequences revealed a close relationship between C. tropicalis and C. parapsilosis, and to a lesser extent C. albicans within the yeast‐like fungi. This group can itself be delineated from predominantly filamentous species. The more distal relationships between Candida (torulopsis) glabrata and C. krusei support previous findings based on small (18S) ribosomal RNA sequence analysis, suggesting a greater degree of evolutionary divergence of these species from the C. albicans group. Among strains of C. albicans we observed conservation of the ITS2 region of the nucleotide level. Conservation was also observed for a more limited number of C. parapsilosis strains. Although the 3′ region of the ITS spacer was species specific, sequence homology was observed in the 5′ end within the albicans/parapsilosis/tropicalis group. Our findings suggest a rapid approach to species identification through the use of non‐conserved regions flanked by highly conserved, functional domains.
The electrophoretic karyotype of two strains of Candida albicans by transverse alternate field electrophoresis reveals higher number of chromosomes ranging from 1 to 3·5 Mb
Authors: Allegrucci Massimo , Lanfaloni Luisa , Bietta Carla , Spaccapelo Roberta , Fioretti Maria C. , Bistoni Francesco 
Published: May 2005

AbstractThe advent of the powerful electrophoretic technique, pulsed field gel electrophoresis, first developed on the yeast Saccharomyces cerevisiæ, has brought a vital impulse to the genetic study on the opportunistic pathogen Candida albicans. We report here on sizing and numbering of Candida chromosomes using transverse alternate field electrophoresis. Our results indicate the occurrence of nine to ten electrophoretic bands (depending on type of Candida strain), that range in approximate size from 1 to 3·5 Mbp, and may account for a higher overall chromosome number, because at least two of these bands appear to be doublets. This number of bands, with smaller size, is considerably higher than previously reported.
Chromatin structure of the yeast FBP1 gene: Transcription‐dependent changes in the regulatory and coding regions
Authors: Del Olmo Marcel Lí , Sogo José M. , Franco Luis , Pérez‐Ortín José E. 
Published: May 2005

AbstractWe have studied the chromatin structure of the Saccharomyces cerevisiae FBP1 gene, which codes for fructose‐1,6‐bisphosphatase. A strong, constitutive, DNase I, micrococcal nuclease and S1 nuclease hypersensitive site is present close to the 3′ end of the coding region. In the repressed state, positioned nucleosomes exist around this site, and subtle changes occur in this nucleosomal organization upon derepression. A DNase I hypersensitive region is located within the promoter between positions −540 and −400 and it extends towards the gene in the derepressed state, leading to an alteration of nucleosomal positioning. Psoralen crosslinking of chromatin, which is used for the first time to study the mobility of restriction fragments from an RNA polymerase II gene, revealed that part of the promoter is nucleosome‐free, in accordance with the results of DNase I digestion. A model is presented that, based on the chromatin structure, puts forward the hypothesis that the promoter UAS is located between − 540 and − 340. Finally, psoralen crosslinking, as well as digestions with micrococcal nuclease or restriction endonucleases suggests that most if not all of the copies of the active FBP1 gene are covered by nucleosomes.
Lysine144 is essential for the catalytic activity of Saccharomyces cerevisiae transaldolase
Authors: Miosga Thomas , Schaaff‐Gerstenschläger Ine , Franken Eva , Zimmermann Friedrich K. 
Published: May 2005

AbstractReplacement of lysine144 by glutamine in the pentose phosphate pathway enzyme transaldolase of Saccharomyces cerevisiae is associated with the complete loss of activity indicating the essential role in catalysis. Neither histidine nor cysteine is important for catalytic activity as proposed for the Candida utilis enzyme. Also we could not find any evidence for a half‐site character of the enzyme as described for transaldolase of C. utilis. Therefore, the reaction mechanisms for the two enzymes are different.
The 5‐aminoimidazole ribonucleotide‐carboxylase structural gene of the methylotrophic yeast Pichia methanolica: Cloning, sequencing and homology analysis
Authors: Hiep Tran Tuan , Kulikov Vladimir N. , Noskov Vladimir N. , Sizonenko Genadij I. , Chernoff Yuri O. , Pavlov Yuri I. 
Published: May 2005

AbstractThe ADE1 gene of the yeast Pichia methanolica encodes phosphoribosyl‐5‐aminoimidazole‐carboxylase (AIRC, EC 4.1.1.21), which is involved in purine biosynthesis. The gene was cloned by complementation of an ade2 mutation in Saccharomyces cerevisiae and a 3077 nucleotide DNA fragment was sequenced. The sequence possessed a single open reading frame, corresponding to a 543 amino acid sequence. The sequence of this putative protein has been compared to the proteins of homologous genes from S. cerevisiae, Schizosaccharomyces pombe, Escherichia coli, chicken and man. The analysis revealed remarkable homology between yeast AIRCs, while for other proteins homology was limited to defined regions.
Formation of the 3′ end of yeast mitochondrial mRNAs occurs by site‐specific cleavage two bases downstream of a conserved dodecamer sequence
Authors: Hofmann Ted Joseph , Min Jingjuan , Zassenhaus Hans Peter 
Published: May 2005

AbstractMitochondrial mRNAs in yeast arise by processing of polygenic primary transcripts at a conserved dodecamer sequence (5′‐AAUAAPyAUUCUU‐3′). Previous results indicated that processing at dodecamer sites interrupted the sequence implying that it functioned primarily as a signal for 3′ end formation of mRNAs. We have determined the precise cleavage site for RNAs processed at the dodecamer sequences associated with the oli1 gene and the ω intron of the 21S rRNA gene. In both cases cleavage occurred two bases downstream of the site. Hydrolysis left the PO4 group attached to the 3′ terminus of the cleavage products. These results demonstrate for the first time that mature mitochondrial mRNAs terminate with an intact dodecamer sequence. In light of the recent identification of a protein complex within mitochondria that binds to RNAs terminating with an intact dodecamer sequence, these results support the idea that the dodecamer sequence functions not only within pre‐mRNAs as a processing site, but within mature mRNAs as well, possibly for the stabilization and/or translation.
Role of O‐acetylhomoserine sulfhydrylase in sulfur amino acid synthesis in various yeasts
Authors: Brzywczy Jerzy , Paszewski Andrzej 
Published: May 2005

AbstractMutants defective in O‐acetylhomoserine sulfhydrylase (OAH‐SHLase) were obtained in five yeast strains representative of different yeast genera: Saccharomyces cerevisiae, Kluyveromyces lactis, Yarrowia lipolytica, Schizosaccharomyces pombe and Trichosporon cutaneum. In vitro, in all five strains, the enzyme also had O‐acetylserine (OAS) sulfhydrylase activity so it is a ‘bifunctional’ OAH/OAS‐SHLase (Yamagata, 1989). The enzyme was only found to be essential in S. cerevisiae (OAH SHLase‐negative mutants are auxotrophs). Its impairment in K. lactis caused a slower growth rate and a decrease of the sulfur amino acid pool. In T. cutaneum only the pool was affected whereas in Y. lipolytica and S. pombe the lesion caused no change in the growth rate nor in the pool. In all strains where OAH SHLase‐negative mutants were prototrophs, a monofunctional OAS sulfhydrylase was detected. The results indicate that OAH SHLase may play different physiological roles in various yeasts.
Extracellular levansucrase of Bacillus subtilis produced in yeast remains in the cell in its precursor form
Authors: Scotti Pier A. , Chambert Régis , Petit‐Glatron Marie‐Françoise 
Published: May 2005

AbstractLevansucrase, a Bacillus subtilis extracellular enzyme, was not secreted in the culture medium when produced in yeast. The protein accumulated inside the cell in its precursor form which represented 0·3% of total proteins. The absence of any post‐translational modifications, such as signal sequence cleavage or addition of N‐linked sugars, indicated that this protein did not enter the reticulum secretion pathway.Direct observation of the cells by confocal laser scanning microscopy showed that levansucrase was associated with the cytoplasmic membrane. Subcellular fractionation experiments revealed that levansucrase precursor form is associated with membranes through weak ionic interactions. The purified precursor displayed the same catalytic properties as levansucrase secreted by B. subtilis. Thus yeast could be used as a source of levansucrase precursor allowing its isolation as a pure form on a milligram scale.