SELECTIVE ABORTION OF TWO NONSISTER NUCLEI IN A DEVELOPING ASCUS OF THE hfd1—1 MUTANT IN SACCHAROMYCES CEREVISIAE

ABSTRACT A recessive mutation, hfd1—1, in strain SOS4 of Saccharomyces cerevisiae leads the mutant cells to produce predominantly two-spored asci. Light microscopical examination of Giemsastained cells revealed no significant differences in the meiotic figures between mutant and wild-type strains. However, only two of the four meiotic products in a developing ascus matured to ascospores in SOS4. Dyad analysis was carried out on an hfd1-1 mutant strain heterozygous for three markers, asp5, gal1 and arg4, which are closely linked to their centromeres, and for his4, which is loosely linked to its centromere. The twospored asci produced by the hfd1—1 mutant segregated dominant (+) and recessive (-) alleles of each marker in a 1:1 ratio; they generally contained one + and one - spore for any given marker. The occurrence of rare dyads with two + or two - spores can be explained quantitatively by recombination between the marker and its centromere. From the results of these cytological and genetical analyses, we infer that, in the mutant strain, one genome set is partitioned to each of the four second-meiotic division poles, but only two nonsister genomes are incorporated into mature spores. Thus, the hfd1—1 mutation in SOS4 blocks incorporation of two nonsister nuclei into mature ascospores, but does not block enclosure of the remaining two nonsister nuclei.

Download Full-text

Barrier Activity in Candida albicans Mediates Pheromone Degradation and Promotes Mating

Eukaryotic Cell ◽

10.1128/ec.00090-07 ◽

2007 ◽

Vol 6 (6) ◽

pp. 907-918 ◽

Cited By ~ 28

Author(s):

Dana Schaefer ◽

Pierre Côte ◽

Malcolm Whiteway ◽

Richard J. Bennett

Keyword(s):

Saccharomyces Cerevisiae ◽

Candida Albicans ◽

Cell Growth ◽

Cell Fusion ◽

Aspartyl Protease ◽

Wild Type ◽

Pheromone Activity ◽

Important Regulator ◽

Mutant Cells ◽

Type Strains

ABSTRACT Mating in Candida albicans and Saccharomyces cerevisiae is regulated by the secretion of peptide pheromones that initiate the mating process. An important regulator of pheromone activity in S. cerevisiae is barrier activity, involving an extracellular aspartyl protease encoded by the BAR1 gene that degrades the alpha pheromone. We have characterized an equivalent barrier activity in C. albicans and demonstrate that the loss of C. albicans BAR1 activity results in opaque a cells exhibiting hypersensitivity to alpha pheromone. Hypersensitivity to pheromone is clearly seen in halo assays; in response to alpha pheromone, a lawn of C. albicans Δbar1 mutant cells produces a marked zone in which cell growth is inhibited, whereas wild-type strains fail to show halo formation. C. albicans mutants lacking BAR1 also exhibit a striking mating defect in a cells, but not in α cells, due to overstimulation of the response to alpha pheromone. The block to mating occurs prior to cell fusion, as very few mating zygotes were observed in mixes of Δbar1 a and α cells. Finally, in a barrier assay using a highly pheromone-sensitive strain, we were able to demonstrate that barrier activity in C. albicans is dependent on Bar1p. These studies reveal that a barrier activity to alpha pheromone exists in C. albicans and that the activity is analogous to that caused by Bar1p in S. cerevisiae.

Download Full-text

Yeast dom34 Mutants Are Defective in Multiple Developmental Pathways and Exhibit Decreased Levels of Polyribosomes

Genetics ◽

10.1093/genetics/149.1.45 ◽

1998 ◽

Vol 149 (1) ◽

pp. 45-56

Author(s):

Luther Davis ◽

JoAnne Engebrecht

Keyword(s):

Saccharomyces Cerevisiae ◽

Cell Cycle ◽

Heterologous Expression ◽

Protein Translation ◽

Developmental Pathways ◽

G1 Phase ◽

Growth Defect ◽

Wild Type ◽

Drosophila Homolog ◽

Mutant Cells

Abstract The DOM34 gene of Saccharomyces cerevisiae is similar togenes found in diverse eukaryotes and archaebacteria. Analysis of dom34 strains shows that progression through the G1 phase of the cell cycle is delayed, mutant cells enter meiosis aberrantly, and their ability to form pseudohyphae is significantly diminished. RPS30A, which encodes ribosomal protein S30, was identified in a screen for high-copy suppressors of the dom34Δ growth defect. dom34Δ mutants display an altered polyribosome profile that is rescued by expression of RPS30A. Taken together, these data indicate that Dom34p functions in protein translation to promote G1 progression and differentiation. A Drosophila homolog of Dom34p, pelota, is required for the proper coordination of meiosis and spermatogenesis. Heterologous expression of pelota in dom34Δ mutants restores wild-type growth and differentiation, suggesting conservation of function between the eukaryotic members of the gene family.

Download Full-text

Influences of Histone Stoichiometry on the Target Site Preference of Retrotransposons Ty1 and Ty2 in Saccharomyces cerevisiae

Genetics ◽

10.1093/genetics/142.3.761 ◽

1996 ◽

Vol 142 (3) ◽

pp. 761-776 ◽

Cited By ~ 2

Author(s):

Lori A Rinckel ◽

David J Garfinkel

Keyword(s):

Saccharomyces Cerevisiae ◽

Promoter Region ◽

Target Site ◽

Site Preference ◽

Wild Type ◽

Histone Proteins ◽

Protein Levels ◽

In The Wild ◽

Type Strains ◽

Orientation Bias

Abstract In Saccharomyces cerevisiae, the target site specificity of the retrotransposon Ty1 appears to involve the Ty integration complex recognizing chromatin structures. To determine whether changes in chromatin structure affect Ty1 and Ty2 target site preference, we analyzed Ty transposition at the CAN1 locus in mutants containing altered levels of histone proteins. A Δhta1-htb1 mutant with decreased levels of H2A and H2B histone proteins showed a pattern of Ty1 and Ty2 insertions at CAN1 that was significantly different from that of both the wild-type and a Δhta2-htb2 mutant, which does not have altered histone protein levels. Altered levels of H2A and H2B proteins disrupted a dramatic orientation bias in the CAN1 promoter region. In the wild-type strains, few Ty1 and Ty2 insertions in the promoter region were oriented opposite to the direction of CAN1 transcription. In the Δhta1-htb1 background, however, numerous Ty1 and Ty2 insertions were in the opposite orientation clustered within the TATA region. This altered insertion pattern does not appear to be due to a bias caused by selecting canavanine resistant isolates in the different HTA1-HTB1 backgrounds. Our results suggest that reduced levels of histone proteins alter Ty target site preference and disrupt an asymmetric Ty insertion pattern.

Download Full-text

CSE4 Genetically Interacts With the Saccharomyces cerevisiae Centromere DNA Elements CDE I and CDE II but Not CDE III: Implications for the Path of the Centromere DNA Around a Cse4p Variant Nucleosome

Genetics ◽

10.1093/genetics/156.3.973 ◽

2000 ◽

Vol 156 (3) ◽

pp. 973-981

Author(s):

Kevin C Keith ◽

Molly Fitzgerald-Hayes

Keyword(s):

Saccharomyces Cerevisiae ◽

Chromosome Loss ◽

Structural Studies ◽

Wild Type ◽

Histone Fold ◽

Histone Fold Domain ◽

Histone H3 Variant ◽

Dna Elements ◽

Mutant Cells ◽

Loss Rates

Abstract Each Saccharomyces cerevisiae chromosome contains a single centromere composed of three conserved DNA elements, CDE I, II, and III. The histone H3 variant, Cse4p, is an essential component of the S. cerevisiae centromere and is thought to replace H3 in specialized nucleosomes at the yeast centromere. To investigate the genetic interactions between Cse4p and centromere DNA, we measured the chromosome loss rates exhibited by cse4 cen3 double-mutant cells that express mutant Cse4 proteins and carry chromosomes containing mutant centromere DNA (cen3). When compared to loss rates for cells carrying the same cen3 DNA mutants but expressing wild-type Cse4p, we found that mutations throughout the Cse4p histone-fold domain caused surprisingly large increases in the loss of chromosomes carrying CDE I or CDE II mutant centromeres, but had no effect on chromosomes with CDE III mutant centromeres. Our genetic evidence is consistent with direct interactions between Cse4p and the CDE I-CDE II region of the centromere DNA. On the basis of these and other results from genetic, biochemical, and structural studies, we propose a model that best describes the path of the centromere DNA around a specialized Cse4p-nucleosome.

Download Full-text

Mild temperature shock alters the transcription of a discrete class of Saccharomyces cerevisiae genes

Molecular and Cellular Biology ◽

10.1128/mcb.3.3.457-465.1983 ◽

1983 ◽

Vol 3 (3) ◽

pp. 457-465

Author(s):

C H Kim ◽

J R Warner

Keyword(s):

Saccharomyces Cerevisiae ◽

Ribosomal Protein ◽

Ribosomal Proteins ◽

Temperature Shift ◽

Restrictive Temperature ◽

Ribosomal Protein Genes ◽

Wild Type ◽

Temperature Shock ◽

Mild Temperature ◽

Mutant Cells

In Saccharomyces cerevisiae the synthesis of ribosomal proteins declines temporarily after a culture has been subjected to a mild temperature shock, i.e., a shift from 23 to 36 degrees C, each of which support growth. Using cloned genes for several S. cerevisiae ribosomal proteins, we found that the changes in the synthesis of ribosomal proteins parallel the changes in the concentration of mRNA of each. The disappearance and reappearance of the mRNA is due to a brief but severe inhibition of the transcription of each of the ribosomal protein genes, although the total transcription of mRNA in the cells is relatively unaffected by the temperature shock. The precisely coordinated response of these genes, which are scattered throughout the genome, suggests that either they or the enzyme which transcribes them has unique properties. In certain S. cerevisiae mutants, the synthesis of ribosomal proteins never recovers from a temperature shift. Yet both the decline and the resumption of transcription of these genes during the 30 min after the temperature shift are indistinguishable from those in wild-type cells. The failure of the mutant cells to grow at the restrictive temperature appears to be due to their inability to process the RNA transcribed from genes which have introns (Rosbash et al., Cell 24:679-686, 1981), a large proportion of which appear to be ribosomal protein genes.

Download Full-text

Farnesyl cysteine C-terminal methyltransferase activity is dependent upon the STE14 gene product in Saccharomyces cerevisiae

Molecular and Cellular Biology ◽

10.1128/mcb.10.10.5071-5076.1990 ◽

1990 ◽

Vol 10 (10) ◽

pp. 5071-5076

Author(s):

C A Hrycyna ◽

S Clarke

Keyword(s):

Saccharomyces Cerevisiae ◽

Cysteine Residue ◽

Peptide Sequence ◽

Direct Result ◽

Wild Type ◽

Methyltransferase Activity ◽

Ras Protein ◽

Carboxyl Methyltransferase ◽

C Terminus ◽

Mutant Cells

Membrane extracts of sterile Saccharomyces cerevisiae strains containing the a-specific ste14 mutation lack a farnesyl cysteine C-terminal carboxyl methyltransferase activity that is present in wild-type a and alpha cells. Other a-specific sterile strains with ste6 and ste16 mutations also have wild-type levels of the farnesyl cysteine carboxyl methyltransferase activity. This enzyme activity, detected by using a synthetic peptide sequence based on the C-terminus of a ras protein, may be responsible not only for the essential methylation of the farnesyl cysteine residue of a mating factor, but also for the methylation of yeast RAS1 and RAS2 proteins and possibly other polypeptides with similar C-terminal structures. We demonstrate that the farnesylation of the cysteine residue in the peptide is required for the methyltransferase activity, suggesting that methyl esterification follows the lipidation reaction in the cell. To show that the loss of methyltransferase activity is a direct result of the ste14 mutation, we transformed ste14 mutant cells with a plasmid complementing the mating defect of this strain and found that active enzyme was produced. Finally, we demonstrated that a similar transformation of cells possessing the wild-type STE14 gene resulted in sixfold overproduction of the enzyme. Although more complicated possibilities cannot be ruled out, these results suggest that STE14 is a candidate for the structural gene for a methyltransferase involved in the formation of isoprenylated cysteine alpha-methyl ester C-terminal structures.

Download Full-text

Protein secretion in Tetrahymena thermophila: characterization of the secretory mutant strain SB281

Journal of Cell Science ◽

10.1242/jcs.78.1.49 ◽

1985 ◽

Vol 78 (1) ◽

pp. 49-65 ◽

Cited By ~ 1

Author(s):

N.J. Maihle ◽

B.H. Satir

Keyword(s):

Plasma Membrane ◽

Mutant Strain ◽

Protein Secretion ◽

Tetrahymena Thermophila ◽

Freeze Fracture ◽

Size Class ◽

Secretory Product ◽

Wild Type ◽

Intramembrane Particle ◽

Mutant Cells

The ciliated protozoon Tetrahymena thermophila contains membrane-bounded secretory organelles termed mucocysts, the release of which has previously been characterized ultrastructurally as a model system for the events occurring during membrane fusion and protein secretion. Recently, a series of secretory mutant strains of Tetrahymena has been isolated following mutagenesis of a parental wild-type strain designated SB210. In this study, the correlates of non-release in one unique mutant strain of this series, designated SB281, are described. SB281 appears to express a diminished (undetectable) level of the major 34000 Mr proteinaceous secretory product of Tetrahymena, as determined by Western immunoblot analysis and indirect immunofluorescence labelling. Thin-section electron-microscopic studies of these cells reveal that they possess no docked or free mature mucocysts. In addition, freeze-fracture electron microscopy demonstrates that an intramembrane particle array termed the rosette, present in the plasma membrane of wild-type cells above sites of docked mucocysts, is absent in the plasma membrane of mutant SB281 cells. A morphometric analysis of intramembrane particles in the plasma membrane of both wild-type and mutant cells indicates that both strains have a similar intramembrane particle density in both leaflets of the the plasma membrane. Although assembled rosettes are missing in the plasma membrane of mutant cells, a 15 nm intramembrane particle size class does exist in the plasma membrane of the mutant, but this size class is significantly reduced in number relative to wild-type.

Download Full-text

Bioreduction of Some Common Carbonylic Compounds Mediated by Yeasts

Zeitschrift für Naturforschung C ◽

10.1515/znc-2010-1-201 ◽

2010 ◽

Vol 65 (1-2) ◽

pp. 1-9 ◽

Cited By ~ 2

Author(s):

Javier Silva ◽

Julio Alarcón ◽

Sergio A. Aguila ◽

Joel B. Alderete

Keyword(s):

Saccharomyces Cerevisiae ◽

Pichia Pastoris ◽

Aqueous Medium ◽

Ethyl Acetoacetate ◽

Enantiomeric Excess ◽

Optically Active ◽

Environmentally Benign ◽

Wild Type ◽

Regeneration System ◽

Type Strains

Bioreduction of several prochiral carbonylic compounds such as acetophenone (1), ethyl acetoacetate (2) and ethyl phenylpropionate (3) to the corresponding optically active secalcohols 1a - 3a was performed using wild-type strains of Pichia pastoris UBB 1500, Rhodotorula sp., and Saccharomyces cerevisiae. The reductions showed moderate to excellent conversion and high enantiomeric excess, in an extremely mild and environmentally benign manner in aqueous medium, using glucose as cofactor regeneration system. The obtained alcohols follow Prelog’s rule, but in the reduction of 1 with P. pastoris UBB 1500 the anti- Prelog enantiopreference was observed

Download Full-text

Mechanisms of sod2 Gene Amplification inSchizosaccharomyces pombe

Molecular Biology of the Cell ◽

10.1091/mbc.11.3.873 ◽

2000 ◽

Vol 11 (3) ◽

pp. 873-886 ◽

Cited By ~ 13

Author(s):

Elizabeth B. Albrecht ◽

Aaron B. Hunyady ◽

George R. Stark ◽

Thomas E. Patterson

Keyword(s):

Gene Amplification ◽

Base Pair ◽

Dna Lesions ◽

The Novel ◽

Wild Type ◽

Resistant Mutants ◽

Chromosome Ii ◽

Chromosome I ◽

Mutant Cells ◽

Type Strains

Gene amplification in eukaryotes plays an important role in drug resistance, tumorigenesis, and evolution. TheSchizosaccharomyces pombe sod2 gene provides a useful model system to analyze this process. sod2 is near the telomere of chromosome I and encodes a plasma membrane Na+(Li+)/H+ antiporter. Whensod2 is amplified, S. pombe survives otherwise lethal concentrations of LiCl, and >90% of the amplifiedsod2 genes are found in 180- and 225-kilobase (kb) linear amplicons. The sequence of the novel joint of the 180-kb amplicon indicates that it is formed by recombination between homologous regions near the telomeres of the long arm of chromosome I and the short arm of chromosome II. The 225-kb amplicon, isolated three times more frequently than the 180-kb amplicon, is a palindrome derived from a region near the telomere of chromosome I. The center of symmetry of this palindrome contains an inverted repeat consisting of two identical 134-base pair sequences separated by a 290-base pair spacer. LiCl-resistant mutants arise 200–600 times more frequently in strains deficient for topoisomerases or DNA ligase activity than in wild-type strains, but the mutant cells contain the same amplicons. These data suggest that amplicon formation may begin with DNA lesions such as breaks. In the case of the 225-kb amplicon, the breaks may lead to a hairpin structure, which is then replicated to form a double-stranded linear amplicon, or to a cruciform structure, which is then resolved to yield the same amplicon.

Download Full-text

The intracellular reserve polysaccharide of Clostridium pasteurianum

Canadian Journal of Microbiology ◽

10.1139/m77-141 ◽

1977 ◽

Vol 23 (8) ◽

pp. 947-953 ◽

Cited By ~ 10

Author(s):

A. G. Darvill ◽

M. A. Hall ◽

J. P. Fish ◽

J. G. Morris

Keyword(s):

Mutant Strain ◽

Minimal Medium ◽

Clostridium Pasteurianum ◽

Wild Type ◽

Methylation Analysis ◽

Glucose Content ◽

Glucose Minimal Medium ◽

Intracellular Polysaccharide ◽

Type Strains ◽

Negative Mutant

An amylopectinlike polysaccharide (granulose) was the only glucan produced in significant quantities by six wild-type strains of Clostridium pasteurianum grown in glucose minimal medium. The intracellular polysaccharide granules laid down before sporulation contained only this amylopectin. No intracellular dextran was discovered in these wild-type strains, nor in a granulose-negative mutant strain of C. pasteurianum possessing an ADP glucose pyrophosphorylase (EC 2.7.7.27) but lacking a granulose synthase (i.e. ADPglucose-α-1,4-glucan glucosyl transferase, EC 2.4.1.21). Furthermore, methylation analysis demonstrated that (1 → 6) linked α-D-glucose units accounted for less than 2% of the entire glucose content of these organisms.

Download Full-text