Multicopy Suppression Screen in the msb3 msb4 Saccharomyces cerevisiae Double Mutant, Affected in Ypt/RabGAP Activity

2005 ◽  
Vol 27 (19) ◽  
pp. 1439-1449 ◽  
Author(s):  
Christophe Dechamps ◽  
Daniel Portetelle ◽  
Micheline Vandenbol
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Double Mutant ◽  
Multicopy Suppression

scholarly journals A Synthetic Interaction between CDC20 and RAD4 in Saccharomyces cerevisiae upon UV Irradiation

2014 ◽  
Vol 2014 ◽  
pp. 1-8
Author(s):  
Bernadette Connors ◽  
Lauren Rochelle ◽  
Asela Roberts ◽  
Graham Howard
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Nucleotide Excision Repair ◽  
Uv Irradiation ◽  
Double Mutant ◽  
Excision Repair ◽  
Strand Break ◽  
Anaphase Promoting Complex ◽  
End Joining ◽  
Ubiquitin Ligase Complex ◽  
Nucleotide Excision

Regulation of DNA repair can be achieved through ubiquitin-mediated degradation of transiently induced proteins. In Saccharomyces cerevisiae, Rad4 is involved in damage recognition during nucleotide excision repair (NER) and, in conjunction with Rad23, recruits other proteins to the site of damage. We identified a synthetic interaction upon UV exposure between Rad4 and Cdc20, a protein that modulates the activity of the anaphase promoting complex (APC/C), a multisubunit E3 ubiquitin ligase complex. The moderately UV sensitive Δrad4 strain became highly sensitive when cdc20-1 was present, and was rescued by overexpression of CDC20. The double mutant is also deficient in elicting RNR3-lacZ transcription upon exposure to UV irradiation or 4-NQO compared with the Δrad4 single mutant. We demonstrate that the Δrad4/cdc20-1 double mutant is defective in double strand break repair by way of a plasmid end-joining assay, indicating that Rad4 acts to ensure that damaged DNA is repaired via a Cdc20-mediated mechanism. This study is the first to present evidence that Cdc20 may play a role in the degradation of proteins involved in nucleotide excision repair.

SPT10 and SPT21 are required for transcription of particular histone genes in Saccharomyces cerevisiae

1994 ◽  
Vol 14 (8) ◽  
pp. 5223-5228
Author(s):  
C Dollard ◽  
S L Ricupero-Hovasse ◽  
G Natsoulis ◽  
J D Boeke ◽  
F Winston
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Differential Expression ◽  
Double Mutant ◽  
The Other ◽  
Histone Genes ◽  
Related Gene ◽  
Histone Proteins ◽  
Histone Locus

The Saccharomyces cerevisiae genome contains four loci that encode histone proteins. Two of these loci, HTA1-HTB1 and HTA2-HTB2, each encode histones H2A and H2B. The other two loci, HHT1-HHF1 and HHT2-HHF2, each encode histones H3 and H4. Because of their redundancy, deletion of any one histone locus does not cause lethality. Previous experiments demonstrated that mutations at one histone locus, HTA1-HTB1, do cause lethality when in conjunction with mutations in the SPT10 gene. SPT10 has been shown to be required for normal levels of transcription of several genes in S. cerevisiae. Motivated by this double-mutant lethality, we have now investigated the interactions of mutations in SPT10 and in a functionally related gene, SPT21, with mutations at each of the four histone loci. These experiments have demonstrated that both SPT10 and SPT21 are required for transcription at two particular histone loci, HTA2-HTB2 and HHF2-HHT2, but not at the other two histone loci. These results suggest that under some conditions, S. cerevisiae may control the level of histone proteins by differential expression of its histone genes.

SAM2 encodes the second methionine S-adenosyl transferase in Saccharomyces cerevisiae: physiology and regulation of both enzymes

1988 ◽  
Vol 8 (12) ◽  
pp. 5132-5139
Author(s):  
D Thomas ◽  
R Rothstein ◽  
N Rosenberg ◽  
Y Surdin-Kerjan
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Gene Disruption ◽  
Double Mutant ◽  
State Level ◽  
Functional Complementation ◽  
The Other ◽  
Steady State Level ◽  
Duplicated Genes ◽  
Double Mutant Strain ◽  
Adomet Synthetase

In Saccharomyces cerevisiae the SAM1 and SAM2 genes encode two distinct forms of S-adenosylmethionine (AdoMet) synthetase. In a previous study we cloned and sequenced the SAM1 gene (D. Thomas and Y. Surdin-Kerjan, J. Biol. Chem. 262:16704-16709, 1987). In this work, the SAM2 gene was isolated by functional complementation of a yeast double-mutant strain, and its identity was ascertained by gene disruption. It has been sequenced and compared with the SAM1 gene. The degree of homology found between the two genes shows that SAM1 and SAM2 are duplicated genes. Using strains disrupted in one or the other SAM gene, we have studied the regulation of their expression by measuring the steady-state level of mRNA after growth under different conditions. The results show that the expression of the two SAM genes is regulated differently, SAM2 being induced by the presence of excess methionine in the growth medium and SAM1 being repressed under the same conditions. The level of mRNA in the parental strain shows that it is not the sum of the levels found in the two disrupted strains. This raises the question of how the two AdoMet synthetases in S. cerevisiae interact to control AdoMet synthesis.

scholarly journals ZDS1 and ZDS2, genes whose products may regulate Cdc42p in Saccharomyces cerevisiae.

1996 ◽  
Vol 16 (10) ◽  
pp. 5264-5275 ◽  
Author(s):  
E Bi ◽  
J R Pringle
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Amino Acids ◽  
Intracellular Signaling ◽  
Double Mutant ◽  
Genome Project ◽  
Yeast Genome ◽  
Glutathione S Transferase ◽  
Gene Encoding ◽  
Biological Phenomena ◽  
Wide Range

A genetic screen for GTPase-activating proteins (GAPs) or other negative regulators of the Rac/Rho family GTPase Cdc42p in Saccharomyces cerevisiae identified ZDS1, a gene encoding a protein of 915 amino acids. Sequence from the yeast genome project identified a homolog, ZDS2, whose predicted product of 942 amino acids is 38% identical in sequence to Zds1p. Zds1p and Zds2p have no detectable homology to known Rho-GAPs or to other known proteins. However, by several assays, it appears that overexpression of either Zds1p or Zds2p decreases the level of Cdc42p activity. Deletion analysis also suggests that Zds1p and Zds2p are at least partially overlapping in function. Deletion of ZDS2 produced no obvious phenotype, and deletion of ZDS1 produced no obvious phenotype other than a mild effect on cell shape. However, the zds1 zds2 double mutant grew slowly with an apparent mitotic delay and produced elongated cells and buds with other evidence of abnormal morphogenesis. A glutathione S-transferase-Zds1p fusion protein that fully complemented the double mutant localized to presumptive bud sites and the tips of small buds. The similarity of this localization to that of Cdc42p suggests that Zds1p may interact directly with Cdc42p. As ZDS1 and ZDS2 have recently been identified also by numerous other groups studying a wide range of biological phenomena, the roles of Cdc42p in intracellular signaling may be more diverse than has previously been appreciated.

scholarly journals The EGP1 gene may be a positive regulator of protein phosphatase type 1 in the growth control of Saccharomyces cerevisiae.

1995 ◽  
Vol 15 (7) ◽  
pp. 3767-3776 ◽  
Author(s):  
N Hisamoto ◽  
D L Frederick ◽  
K Sugimoto ◽  
K Tatchell ◽  
K Matsumoto
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Growth ◽  
Protein Phosphatase ◽  
Double Mutant ◽  
Growth Control ◽  
Restrictive Temperature ◽  
Extra Copy ◽  
Additional Gene ◽  
Positive Modulator

The Saccharomyces cerevisiae GLC7 gene encodes the catalytic subunit of type 1 protein phosphatase (PP1) and is required for cell growth. A cold-sensitive glc7 mutant (glc7Y170) arrests in G2/M but remains viable at the restrictive temperature. In an effort to identify additional gene products that function in concert with PP1 to regulate growth, we isolated a mutation (gpp1) that exacerbated the growth phenotype of the glc7Y170 mutation, resulting in rapid death of the double mutant at the nonpermissive temperature. We identified an additional gene, EGP1, as an extra-copy suppressor of the glc7Y170 gpp1-1 double mutant. The nucleotide sequence of EGP1 predicts a leucine-rich repeat protein that is similar to Sds22, a protein from the fission yeast Schizosaccharomyces pombe that positively modulates PP1. EGP1 is essential for cell growth but becomes dispensable upon overexpression of the GLC7 gene. Egp1 and PP1 directly interact, as assayed by coimmunoprecipitation. These results suggest that Egp1 functions as a positive modulator of PP1 in the growth control of S. cerevisiae.

scholarly journals Bud8p and Bud9p, Proteins That May Mark the Sites for Bipolar Budding in Yeast

2001 ◽  
Vol 12 (8) ◽  
pp. 2497-2518 ◽  
Author(s):  
Heidi A. Harkins ◽  
Nicolas Pagé ◽  
Laura R. Schenkman ◽  
Claudio De Virgilio ◽  
Sidney Shaw ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Double Mutant ◽  
Previous Analysis ◽  
Integral Membrane Proteins ◽  
Cell Cortex ◽  
Daughter Cells ◽  
Distal Pole ◽  
Bud Position ◽  
Bud Site ◽  
Hydrophilic Domain

The bipolar budding pattern of a /α Saccharomyces cerevisiae cells appears to depend on persistent spatial markers in the cell cortex at the two poles of the cell. Previous analysis of mutants with specific defects in bipolar budding identifiedBUD8 and BUD9 as potentially encoding components of the markers at the poles distal and proximal to the birth scar, respectively. Further genetic analysis reported here supports this hypothesis. Mutants deleted for BUD8 orBUD9 grow normally but bud exclusively from the proximal and distal poles, respectively, and the double-mutant phenotype suggests that the bipolar budding pathway has been totally disabled. Moreover, overexpression of these genes can cause either an increased bias for budding at the distal (BUD8) or proximal (BUD9) pole or a randomization of bud position, depending on the level of expression. The structures and localizations of Bud8p and Bud9p are also consistent with their postulated roles as cortical markers. Both proteins appear to be integral membrane proteins of the plasma membrane, and they have very similar overall structures, with long N-terminal domains that are both N- andO-glycosylated followed by a pair of putative transmembrane domains surrounding a short hydrophilic domain that is presumably cytoplasmic. The putative transmembrane and cytoplasmic domains of the two proteins are very similar in sequence. When Bud8p and Bud9p were localized by immunofluorescence and tagging with GFP, each protein was found predominantly in the expected location, with Bud8p at presumptive bud sites, bud tips, and the distal poles of daughter cells and Bud9p at the necks of large-budded cells and the proximal poles of daughter cells. Bud8p localized approximately normally in several mutants in which daughter cells are competent to form their first buds at the distal pole, but it was not detected in abni1 mutant, in which such distal-pole budding is lost. Surprisingly, Bud8p localization to the presumptive bud site and bud tip also depends on actin but is independent of the septins.

scholarly journals The genetic control of direct-repeat recombination in Saccharomyces: the effect of rad52 and rad1 on mitotic recombination at GAL10, a transcriptionally regulated gene.

Genetics ◽  
1989 ◽  
Vol 123 (4) ◽  
pp. 725-738 ◽  
Author(s):  
B J Thomas ◽  
R Rothstein
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Genetic Control ◽  
Plasmid Dna ◽  
Double Mutant ◽  
Direct Repeat ◽  
Single Mutant ◽  
Plasmid Loss ◽  
Mutant Strains ◽  
Recombination Pathway

Abstract We have previously shown direct-repeat recombination events leading to loss of a plasmid integrated at the GAL10 locus in Saccharomyces cerevisiae are stimulated by transcription of the region. We have examined the role of two recombination- and repair-defective mutations, rad1 and rad52, on direct repeat recombination in transcriptionally active and inactive sequences. We show that the RAD52 gene is required for transcription-stimulated recombination events leading to loss of the integrated plasmid. Similarly, Gal+ events between the duplicated repeats that retain the integrated plasmid DNA (Gal+ Ura+ replacement events) are reduced 20-fold in the rad52 mutant in sequences that are constitutively expressed. In contrast, in sequences that are not expressed, the rad52 mutation reduces plasmid loss events by only twofold and Gal+ Ura+ replacements by fourfold. We also observe an increase in disome-associated plasmid loss events in the rad52 mutant, indicative of chromosome gain. This event is not affected by expression of the region. Plasmid loss events in rad1 mutant strains are reduced only twofold in transcriptionally active sequences and are not affected in sequences that are repressed. However, the rad1 and rad52 double mutant shows a decrease in plasmid loss events greater than the sum of the decreases in the rates of this event displayed by either single mutant in both constitutive and repressed DNA, indicating a synergistic interaction between these two genes. The synergism is limited to recombination since the rad1 rad52 double mutant is no more sensitive when compared with either single mutant in its ability to survive radiation damage. Finally, the recombination pathway that remains in the double mutant is positively affected by transcription of the region.

scholarly journals Altered Na+ and Li+Homeostasis in Saccharomyces cerevisiae Cells Expressing the Bacterial Cation Antiporter NhaA

1998 ◽  
Vol 180 (12) ◽  
pp. 3131-3136 ◽  
Author(s):  
Roc Ros ◽  
Consuelo Montesinos ◽  
Abraham Rimon ◽  
Etana Padan ◽  
Ramón Serrano
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Plasma Membrane ◽  
Double Mutant ◽  
Vacuolar Membrane ◽  
Intracellular Sodium ◽  
Intracellular Pool ◽  
Sodium Accumulation ◽  
Sodium Tolerance ◽  
Yeast Mutants ◽  
In Cells

ABSTRACT The bacterial Na+(Li+)/H+antiporter NhaA has been expressed in the yeast Saccharomyces cerevisiae. NhaA was present in both the plasma membrane and internal membranes, and it conferred lithium but not sodium tolerance. In cells containing the yeast Ena1-4 (Na+, Li+) extrusion ATPase, the extra lithium tolerance conferred by NhaA was dependent on a functional vacuolar H+ ATPase and correlated with an increase of lithium in an intracellular pool which exhibited slow efflux of cations. In yeast mutants without (Na+, Li+) ATPase, lithium tolerance conferred by NhaA was not dependent on a functional vacuolar H+ ATPase and correlated with a decrease of intracellular lithium. NhaA was able to confer sodium tolerance and to decrease intracellular sodium accumulation in a double mutant devoid of both plasma membrane (Na+, Li+) ATPase and vacuolar H+ ATPase. These results indicate that the bacterial antiporter NhaA expressed in yeast is functional at both the plasma membrane and the vacuolar membrane. The phenotypes conferred by its expression depend on the functionality of plasma membrane (Na+, Li+) ATPase and vacuolar H+ ATPase.

STUDY OF SACCHAROMYCES CEREVISIAE YEAST STRAIN SENSITIVITY WITH DOUBLE MUTANT RAD2∆HSM3∆ TO METHYLMETHANESULPHONATE

2020 ◽  
pp. 325-328
Author(s):  
E. Alekseeva ◽  
V. Korolev
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Yeast Strain ◽  
Double Mutant ◽  
Methyl Methanesulfonate ◽  
Strain Sensitivity ◽  
Spontaneous Mutagenesis

The sensitivity of the yeast strain Saccharomyces cerevisiae with the double mutant rad2∆hsm3∆ to genotoxicants was studied using methyl methanesulfonate as an example, as well as its influence on the possibility of spontaneous mutagenesis, and further use of the strain as a test for determining genotoxicants in the environment.

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