bluuebunny/crossref_metadata_2025_split huggingface dataset:
In the budding yeast Kluyveromyces marxianus, adenylate cyclase is regulated by Ras protein(s) in vitro
Authors: Verzotti Enrico , Geymonat Marco , Valetti Francesca , Lanzetti Letizia , Giunta Carlo 
Published: May 2005

AbstractThe presence of adenylate cyclase activity was first demonstrated in membrane fractions from the budding yeast Kluyveromyces marxianus. The enzyme showed a Mn2+‐ and Mg2+‐dependent activity, with optimal pH at around 6 as observed in other yeast species. As in Saccharomyces cerevisiae, where adenylate cyclase is regulated by RAS1 and RAS2, we detected a guanyl nucleotide‐dependent activity. Interestingly Y13–259 monoclonal antibody, raised against mammalian p21Ha‐ras, inhibited Mg2+ plus GTP‐γ‐S‐dependent cAMP production, suggesting that the GTP binding proteins involved in adenylate cyclase regulation could be Ras proteins. The same antibody recognized on Western blot and immunoprecipitated a 40 kDa polypeptide from K. marxianus crude membranes. This polypeptide was not detected by an anti‐RAS2 polyclonal antibody raised against S. cerevisiae RAS2 protein, suggesting that Ras proteins from the two species could be structurally different.
Predominant localization of non‐specific lipid‐transfer protein of the yeast Candida tropicalis in the matrix of peroxisomes
Authors: Tan Hironobu , Bun‐Ya Masanori , Hirata Aiko , Kamiryo Tatsuyuki 
Published: May 2005

AbstractPXP‐18 is a 14‐kDa major peroxisomal protein of the yeast Candida tropicalis and a homologue of the non‐specific lipid‐transfer protein (nsLTP) of mammals. Mammalian nsLTP is thought to facilitate the contact of membranes, to stimulate lipid‐transfer between them. If PXP‐18 functions like nsLTP, it must be present on organelle membranes. Immunoelectron microscopy of C. tropicalis cells indicated that gold particles, which visualized PXP‐18, localized exclusively in the matrix of peroxisomes. Subcellular fractionation followed by Western blotting revealed the association of PXP‐18 with peroxisomes in C. tropicalis cells. An enzyme‐linked immunosorbent assay revealed that almost all the PXP‐18 associated with peroxisomes was detectable after the solubilization of the organelle but not before, implying the predominance of PXP‐18 inside peroxisomes. This differential assay was applied to the intracellular import of the intact and truncated PXP‐18s expressed in Saccharomyces cerevisiae cells. Most of the intact PXP‐18 was shown to be imported into the matrix of host‐cell peroxisomes, whereas the truncated PXP‐18, which lacked the C‐terminal tripeptide Pro‐Lys‐Leu, no longer targeted peroxisomes. These results are consistent with the view that PXP‐18 is the matrix protein of peroxisomes and must function in a system other than that of lipid transfer.
II. Yeast sequencing reports. The nucleotide sequence of TTP1, a gene encoding a predicted type II membrane protein
Authors: Romero Pedro A. , Athanassiadis Ariadni , Lussier Marc , Herscovics Annette 
Published: May 2005

AbstractThe DNA sequence of a 2967 bp fragment located near the centromere of chromosome II, between the CEN2 and FUR4 genes, was determined. The segment contains a new open reading frame of 1794 bp. The product encoded by the gene, designated TTP1, is a predicted type II membrane protein of 597 amino acid residues with a short cytoplasmic NH2‐terminus, a membrane‐spanning region and a large COOH‐terminal region containing three potential N‐glycosylation sites. Gene disruption indicated that TTP1 is not essential for cell growth. The sequence has been deposited in the GenBank data library under Accession Number U05211.
Identification of a set of yeast genes coding for a novel family of putative ATPases with high similarity to constituents of the 26S protease complex
Authors: Schnall Ralf , Mannhaupt Gertrud , Stucka Rolf , Tauer Reimund , Ehnle Susanne , Schwarzlose Christa , Vetter Irene , Feldmann Horst 
Published: May 2005

AbstractThere is accumulating evidence for a large, highly conserved gene family of putative ATPases. We have identified 12 different members of this novel gene family (the YTA family) in yeast and determined the nucleotide sequences of nine of these genes. All of the putative gene products are characterized by the presence of a highly conserved domain of 300 amino acids containing specialized forms of the A and B boxes of ATPases. YTA1, YTA2, YTA3 and YTA5 exhibit significant similarity to proteins involved in human immunodeficiency virus Tat‐mediated gene expression but more significantly to subunits of the human 26S proteasome. YTA1 and YTA2 are essential genes in yeast. Remarkably, the cDNA of human TBP‐1 can compensate for the loss of YTA1. Preliminary experiments indicate that YTA1 is a component of the 26S protease complex from yeast. Our findings lead us to propose that YTA1, YTA2, YTA3 and YTA5 function as regulatory subunits of the yeast 26S proteasome. YTA10, YTA11 and YTA12 share significant homology with the Escherichia coli FtsH protein, and together with YTA4 and YTA6 may constitute a separate subclass within this family of putative ATPases.
Purification and immunolocalization of the peroxisomal 3‐oxoacyl‐coA thiolase from Saccharomyces cerevisiae
Authors: Erdmann Ralf , Kunau Wolf‐Hubert 
Published: May 2005

AbstractA molecular understanding of peroxisome biogenesis depends upon the analysis of peroxisomal proteins. Here we describe the isolation of the 3‐oxoacyl‐CoA thiolase of the peroxisomal β‐oxidation system from Saccharomyces cerevisiae as a dimer of identical subunits, each with a molecular mass of 45 kDa. Monospecific polyclonal antibodies were raised against the purified enzyme, and its peroxisomal origin was demonstrated by immunoblotting of subcellular fractions as well as by immunogold labelling. We also show that these antibodies could be suitable for an immunofluorescence microscopy screening of yeast mutants affected in peroxisome assembly.
The in vivo effects of ethidium bromide on mitochondrial and ribosomal DNA in Candida parapsilosis
Authors: Maleszka R. 
Published: May 2005

AbstractThe ability of Candida parapsilosis to grow in the presence of high levels of ethidium bromide (EB) has been explored to study the effects of this intercalating dye on DNA in vivo. By employing confocal microscopy we have determined that EB penetrates the cellular membranes and binds rapidly to the nucleolus, whereas mitochondrial DNA becomes stained after a longer exposure to this dye. No detectable staining of the nucleus has been detected under these conditions.Electrophoretic studies of both undigested and restricted DNAs confirm that the nuclear DNA is unaffected by high levels of EB, with the exception of the rDNA‐bearing chromosome that undergoes significant structural alterations in the presence of EB. Moreover, the hybridization signal with the rDNA probe is proportionally reduced in samples obtained from cultures grown in the presence of EB, suggesting that the average copy number of rRNA genes in these cultures may be affected.In striking contrast to other fungal species, the linear organelle genome in C. parapsilosis retains its structural and functional integrity in the presence of high concentrations of EB.
IV. Yeast sequencing reports. Nucleotide sequence of the SAC2 gene of Saccharomyces cerevisiae
Authors: Kölling Ralf , Lee Angela , Chen Ellson Y. , Botstein David 
Published: May 2005

AbstractA temperature‐sensitive mutation (act1‐1) in the essential actin gene of Saccharomyces cerevisiae can be suppressed by mutations in the SAC2 gene. A cloned genomic DNA fragment that complements the cold‐sensitive growth phenotype associated with such a suppressor mutation (sac2‐1) was sequenced. The fragment contained an open reading frame that encodes a 641 amino acid predicted hydrophilic protein with a molecular weight of 74 445. No sequences with significant similarity to SAC2 were found in the GenBank and EMBL databases. A SAC2 disruption mutation was constructed which had phenotypes similar to the sac2‐1 point mutation. A haploid SAC2 disruption strain failed to grow at low temperature and the disruption allele suppressed the temperature‐sensitive act1‐1 growth defect. The suppression phenotype was dependent on the strain background. The SAC2 sequence has been submitted to the EMBL data library (Accession Number Z29988).
II. Yeast sequencing reports. The sequence of 12·5 kb from the right arm of chromosome II predicts a new N‐terminal sequence for the IRA1 protein and reveals two new genes, one of which is a DEAD‐box helicase
Authors: Zagulski M. , Bécam A‐M , Grzybowska E. , Lacroute F. , Migdalski A. , Slonimski P. P. , Sokolowska B. , Herbert C. J. 
Published: May 2005

AbstractWe have determined the complete nucleotide sequence of a 12·5 kb segment from the right arm of chromosome II carried by the cosmid α20. The sequence encodes the 5′ end of the IRA1 gene. Two complete new open reading frames and the 3′ non‐coding region of the SUP1 (SUP45) gene. A comparison of our sequence with the data bank reveals a 154 amino acid extension at the N‐terminus of Ira1p compared to the previously predicted sequence. According to the 11th edition of the Saccharomyces cerevisiae genetic map, our sequence should encode the MAK5 gene, which is necessary for the maintenance of dsRNA killer plasmids. One of the two new open reading frames, YBR1119, is predicted to encode an RNA helicase, thus YBR1119 may correspond to the MAK5 gene. The sequence has been deposited in the EMBL data library under Accession Number X78937.
A genetic screen to isolate genes regulated by the yeast CCAAT‐box binding protein Hap2p
Authors: Dang Van‐Dinh , Valens Michèle , Bolotin‐Fukuhara Monique , Daignan‐Fornier Bertrand 
Published: May 2005

AbstractWe have developed a screening method to isolate yeast genes regulated by a specific transcription activator. The screen is based on the use of expression libraries in which the lacZ reporter gene is placed under control of yeast regulatory elements. Two partially representative libraries, constructed by different methods, were used to isolate genes regulated by the yeast CCAAT‐box binding protein Hap2p. Among 26 fusions shown to be regulated by Hap2p only CYT1 was known to be regulated by this activator. Sequence analysis revealed that most of the remaining regulated fusions are in new yeast genes, while some are in previously characterized yeast genes (PTP1, RPM2, SDH1). Optimal expression of these three genes also requires Hap3p and Hap4p and is regulated by carbon source. Hap2p was known to regulate expression of genes involved in Krebs cycle, electron transport and heme biosynthesis. Our results suggest that Hap2p could play a more general role by regulating other mitochondrial processes such as protein import and phosphate transport (PTP1) or maturation of mitochondrial tRNAs (RPM2). Among the remaining regulated fusions, two of them correspond to open reading frames (ORFs) on chromosomes III and XI whose nucleotide sequences have been entirely determined. The use of this approach to functionally analyse ORFs of unknown function is discussed.
XIV. Yeast sequencing reports. Sequence of MKT1, needed for propagation of M2 satellite dsRNA of the L‐A virus of Saccharomyces cerevisiae
Authors: Vermut Michael , Widner William R. , Dinman Jonathan D. , Wickner Reed B. 
Published: May 2005

AbstractMKT1 is required for maintenance of K2 above 30°C in strains with the L‐A‐HN variant of the L‐A double‐stranded RNA virus of Saccharomyces cerevisiae. We report that MKT1 encodes a 92 979 Da protein with serine‐rich regions and the retroviral protease signature, DTG, but with no substantial homology to proteins presently in the databases. This sequence is available from GenBank under Accession Number U09129.
X. Yeast sequencing reports. Sequence and function analysis of a 9·74 kb fragment of Saccharomyces cerevisiae chromosome X including the BCK1 gene
Authors: Miosga T. , Boles E. , Schaaff‐Gerstenschläger I. , Schmitt S. , Zimmermann F. K. 
Published: May 2005

AbstractIn the framework of the European BIOTECH project for sequencing the Saccharomyces cerevisiae genome, we have determined the nucleotide sequence of the cosmid clone 233 provided by F. Galibert (Rennes Cedex, France). We present here 9743 base pairs of sequence derived from the left arm of chromosome X. This sequence reveals three new open reading frames and includes the published sequence (5′ end and open reading frame) of the gene BCK1/SLK1/SSP31 also identified as ORFAA. Deletion mutants of two earlier unknown open reading frames J0840 and J0904 are viable and the open reading frame J0902 is essential for yeast growth. The sequence has been entered in the EMBL data library under accession number X77923.
V. Yeast sequencing reports. UBP5 encodes a putative yeast ubiquitin‐specific protease that is related to the human Tre‐2 oncogene product
Authors: Xiao Wei , Fontanie Treena , Tang Min 
Published: May 2005

AbstractA gene from chromosome V of the yeast Saccharomyces cerevisiae has been cloned and sequenced. The deduced amino acid sequence encoded by this gene is similar to several ubiquitin‐specific proteases from yeast, especially at the highly conserved domain. It is thus named UBP5. UBP5 is also closely related to the human Tre‐2 and the mouse Unp oncogene products. This study adds a new member to the ubiquitin protease family and suggests that alteration of ubiquitin protease activity may result in cancer in mammals. However, disruption of the UBP5 gene in a haploid strain did not result in a noticeable phenotypic alteration. The sequence has been deposited in the GenBank data library under Accession Number U10082.
XVI. Yeast sequencing reports. DNA sequence analysis of a 10·4 kbp region on the right arm of yeast chromosome XVI positions GPH1 and SGV1 adjacent to KRE6, and identifies two novel tRNA genes
Authors: Roemer Terry , Fortin Nathalie , Bussey Howard 
Published: May 2005

AbstractDetermination of DNA sequence in the KRE6 region of the Saccharomyces cerevisiae genome completes a 10·4 kbp section on the extreme right arm of chromosome XVI. This segment contains two additional genes, GPH1 and SGV1 (Hwang et al., 1989; Irei et al., 1991) previously assigned physically to chromosome XVI, as well as a new tRNAGly gene, and a novel tRNAAla gene which is flanked by a sigma element. The complete 10·4 kbp DNA sequence has been entered in GenBank with accession number L33835.
IIX. Yeast mapping reports. ATP1 and ATP2, the F1F0‐ATPase α and β subunit genes of Saccharomyces cerevisiae, are respectively located on chromosomes II and X
Authors: Takeda Masaharu , Okushiba Takahito , Hayashida Takeya , Gunge Norio 
Published: May 2005

AbstractSouthern blot analysis showed that ATP1 and ATP2 map on chromosomes II and X, respectively. Physical mapping of ATP1 and ATP2 by chromosome fragmentation showed that ATP1 is at the left end of chromosome II and ATP2 is at the right end of chromosome X. Both are located close to telomere sequences of each chromosome; ATP1 and ATP2 being approximately 30 kb and 85 kb from the respective telomeres.
Increase in copy number of an integrated vector during continuous culture of Hansenula polymorpha expressing functional human haemoglobin
Authors: Gilbert S. C. , van Urk H. , Greenfield A. J. , McAvoy M. J. , Denton K. A. , Coghlan D. , Jones G. D. , Mead D. J. 
Published: May 2005

AbstractRecombinant human haemoglobin A (rHbA) was produced by a leucine‐requiring strain of Hansenula polymorpha which had been transformed with an integration vector containing the Saccharomyces cerevisiae LEU2 gene and cDNAs for the expression of α and β globin each driven by the H. polymorpha MOX promoter. After 40 generations in a chemostat it was found that the integrated vector had become amplified in the host strain. In some cases this led to an increase in LEU2 gene dosage, but a loss of globin expression cassettes. In other cases the globin gene dosage also increased. These changes coincided with an increase in rHbA production in the culture, which was reversed when the dilution rate was increased. Isolates from a chemostat culture producing elevated levels of rHbA were grown in fed‐batch fermentations, resulting in higher productivities than when inoculated with the parent strain. The rHbA produced was purified and characterized. Oxygen binding studies and electrospray mass spectrometry showed that the rHbA had been processed and assembled correctly, and behaved as a fully functional co‐operative tetramer.
Effect of site‐directed mutagenesis of conserved lysine residues upon Pas1 protein function in peroxisome biogenesis
Authors: Krause Thomas , Kunau Wolf‐H. , Erdmann Ralf 
Published: May 2005

AbstractThe Pas1 protein (Pas1p) is required for peroxisome biogenesis in Saccharomyces cerevisiae and contains two putative ATP‐binding sites, each within a domain which is conserved among members of the recently characterized AAA‐family. To elucidate whether both putative ATP‐binding sites are essential for Pas1p function, lysine467 of the first and lysine744 of the second putative ATP‐binding site were each changed to glutamate by site‐directed mutagenesis. While replacement of lysine744 abolished the function of the Pas1 protein in peroxisome biogenesis, replacement of lysine467 had no obvious effect.
XIV. Yeast sequencing reports. A 21·7 kb DNA segment on the left arm of yeast chromosome XIV carries WHI3, GCR2, SPX18, SPX19, an homologue to the heat shock gene SSB1 and 8 new open reading frames of unknown function
Authors: Jonniaux Jean Luc , Coster Francoise , Purnelle Benedicte , Goffeau Andre 
Published: May 2005

AbstractWe report the amino acid sequence of 13 open reading frames (ORF > 299 bp) located on a 21·7 kb DNA segment from the left arm of chromosome XIV of Saccharomyces cerevisiae. Five open reading frames had been entirely or partially sequenced previously: WHI3, GCR2, SPX19, SPX18 and a heat shock gene similar to SSB1. The products of 8 other ORFs are new putative proteins among which N1394 is probably a membrane protein. N1346 contains a leucine zipper pattern and the corresponding ORF presents an HAP (global regulator of respiratory genes) upstream activating sequence in the promoting region. N1386 shares homologies with the DNA structure‐specific recognition protein family SSRPs and the corresponding ORF is preceded by an MCB (MluI cell cycle box) upstream activating factor. The sequence has been deposited in the EMBL data library under Accession Number X78898.
II. Yeast sequencing reports. Analysis of a 17·4 kb DNA segment of yeast chromosome II encompassing the ribosomal protein L19 as well as proteins with homologies to components of the hnRNP and snRNP complexes and to the human proliferation‐associated p120 antigen
Authors: van Dyck Luc , Jonniaux Jean‐Luc , de Melo Barreiros Tania , Goffeau Andre , Kleine Karl 
Published: May 2005

AbstractWe report the nucleotide sequence of a 17·4 kb DNA segment from the left arm of Saccharomyces cerevisiae chromosome II. This sequence contains 12 open reading frames (ORFs) longer than 300 bp and a putative autonomously replicating sequence (ARS). The ORF YBL0418 contains the KH motif present in several nucleic acid‐binding proteins and shares homologies with the mouse X protein of the heterogeneous nuclear ribonucleo‐protein (hnRNP) complexes involved in pre‐mRNA processing. YBL0424 is the yeast member of the ribosomal protein L19 (YL14) family. YBL0425 is related to the D1 core polypeptide of the small nuclear ribonucleoprotein (snRNP) particles involved in the splicing of introns. YBL0437 is a putative homologue of the human protein p120, one of the major antigens associated with malignant tumours. Mcm2, a protein important for ARS activity, as well as Aac2, one of the three isoforms of the mitochondrial ATP/ADP carrier, were previously described (Yan et al., 1991; Lawson and Douglas, 1988). Four ORFs show no homology or particular features that could help to assess their functions. The last ORFs are not likely to be expressed for they are localized on the complementary strand of longer ORFs. The sequence has been submitted to the EMBL data library under Accession Number X77291.
The IFH1 gene product interacts with a fork head protein in Saccharomyces cerevisiae
Authors: Cherel Isabelle , Thuriaux Pierre 
Published: May 2005

AbstractFHL1 encodes a polypeptide closely related to the fork head protein family of transcriptional activators. Deleting this gene leads to a slow‐growth phenotype with impaired rRNA maturation. IFH1 (located on chromosome IV) was isolated as a dosage‐dependent suppressor partially correcting the growth defect of the fhl1 deletion. It codes for a highly hydrophilic protein with a predicted molecular weight of 122 kDa and a pI of 4·8, that is very rich in charged residues (mostly acidic) but otherwise unrelated to any known protein. Carboxy‐terminal deletions removing the last third of the protein lead to a leaky growth phenotype with impaired rRNA maturation, as in the case of the fhl1 deletion. A full deletion of IFH1 is lethal, but growth was restored in a strain deleted for both IFH1 and FHL1. Thus, Ifh1p is essential for growth, but only in the presence of a functional Fhp1p protein. Conversely, its overexpression by increased gene dosage partially compensates for the genetic inactivation of FHl1p. These data suggest a direct interaction between the Fhl1p and Ifh1p proteins, and are consistent with a model where Fhl1p is converted from a transcriptional repressor to an activator on binding of Ifh1p. The sequence has been deposited in the EMBL data library under Accession Number Z29488.
Studies on the transformation of intact yeast cells by the LiAc/SS‐DNA/PEG procedure
Authors: Gietz R. Daniel , Schiestl Robert H. , Willems Andrew R. , Woods Robin A. 
Published: May 2005

AbstractAn improved lithium acetate (LiAc)/single‐stranded DNA (SS‐DNA)/polyethylene glycol (PEG) protocol which yields >1 × 106 transformants/μg plasmid DNA and the original protocol described by Schiestl and Gietz (1989) were used to investigate aspects of the mechanism of LiAc/SS‐DNA/PEG transformation. The highest transformation efficiency was observed when 1 × 108 cells were transformed with 100 ng plasmid DNA in the presence of 50 μg SS carrier DNA. The yield of transformants increased linearly up to 5 μg plasmid per transformation. A 20‐min heat shock at 42°C was necessary for maximal yields. PEG was found to deposit both carrier DNA and plasmid DNA onto cells. SS carrier DNA bound more effectively to the cells and caused tighter binding of 32P‐labelled plasmid DNA than did double‐stranded (DS) carrier. The LiAc/SS‐DNA/PEG transformation method did not result in cell fusion. DS carrier DNA competed with DS vector DNA in the transformation reaction. SS plasmid DNA transformed cells poorly in combination with both SS and DS carrier DNA. The LiAc/SS‐DNA/PEG method was shown to be more effective than other treatments known to make cells transformable. A model for the mechanism of transformation by the LiAc/SS‐DNA/PEG method is discussed.